3^rd International Conference on Genetic and Protein Engineering(10 Plenary Forums - 1 Event) November 08-09, 2017 Las Vegas, USA

Biography:

Ron L Martin has received his BS and MS degrees in Food Science and Nutrition from Chapman University in Orange, California. He has worked for more than 35 years in the nutraceutical and food industries, including Hunt-Wesson/ConAgra, Plus Products, the William T Thompson Co., and the Nutrilite division of Alticor, where he served as a Senior Research Scientist in Nutrilite's New Concepts group. He has been a Professional Member of the Institute of Food Technologists (IFT) and is the President of Nutrigenetics Unlimited, Inc., which he founded in 2007. He is also one of the Founding Members of the International Society of Nutrigenetics/Nutrigenomics

Abstract:

Examples of the emerging gene expression evidence-base will be described, along with online tools for increasing both awareness and utility of the increasingly actionable information. This can be helpful to the entire spectrum of potential users, including students and other members of the public. Such tools are increasingly important as the body of literature continues to expand rapidly, making it progressively more difficult to identify and manage the evidence-base for making more fully informed choices. Online resources will be described, including the usefulness of standardized terminology which allows creation of subtopic listings for any given topic or for any given combination of topics – including for genes and gene variants. Beyond diagnosis and treatment alone, such approaches also allow identification and exploration of prevention opportunities, and as well as optimizations for both physical and mental health. Gene-environment examples include nutrition, pharmaceuticals, pollution, lifestyle, social environment, etc. Because nutrition applies to everyone without exception, it can become a useful introductory "archetype" for promoting greater engagement and greater genetics/health literacy.

Biography:

Yuri L Lyubchenko is a Professor of Pharmaceutical Sciences at the University of Nebraska Medical Center, USA. His research focuses on understanding fundamental mechanisms underlying health and disease, which are keys to developing new and more effective diagnostics and medications. This primarily basic research allows him not only to identify new drug targets for small molecule drugs, it also leads to development of the nanotools and methods to discover novel approaches for diagnostic, treatment and disease prevention and to more rapidly determine their effi cacy at the molecular level.

Abstract:

Statement of the Problem: Increasing evidence suggests that the self-assembly of amyloid β(Aβ) protein underlies the early onset of Alzheimer’s disease (AD). Given that small Aβ nano-assemblies (oligomers) are the most neurotoxic species, they have become the major target for the development of treatments and early diagnostic tools for AD. However, advances surrounding this are blocked by the lack of structure intrinsic to Aβ oligomers, as they are transient states of Aβ aggregation kinetics; making traditional structural approaches non-amenable. We have previously developed single-molecule approaches capable of probing of aμo dimers. Here, we extended our approaches to higher order oligomers. We hypothesized that the folding pattern of amyloid protein defines the aggregation pathway. 

Methodology & Theoretical Orientation: In this study, we tested this hypothesis using Aβ( 14-23) peptide in linear form and its tandem assembled in the hairpin-type shape. We combined two experimental approaches and molecular dynamics simulations to characterize molecular interactions and the stability of complexes between Aβ (14-23) hairpin and Aβ (14-23) monomer, as well as the interactions between two hairpins.

Findings: The lifetime measurements demonstrate that the Aβ (14-23) hairpin and a Aβ(14-23) monomer assemble in a very stable complex when compared with homologous ensembles. We measured the strength of hairpin-hairpin and hairpin-monomer interactions which demonstrated that the hairpin-monomer interaction is stronger compared with the hairpin-hairpin assembly; data that is fully in line with the lifetime measurements. Aggregation studies demonstrate that the Aβ(14-23) monomer formed fibrils and the hairpin formed spherical structures. However, their mixture formed neither fibrils nor spherical structures, but rather disk shaped nanostructures.

Conclusion & Significance: Overall, our study provides new insight into the role of the monomer structure on the self-assembly process that contributes to the formation of disease aggregates. Importantly, the developed experimental approaches and validation approaches for computational analyses are not limited to amyloid proteins, but can also be applied to other molecular systems.

Biography:

Barry L Stoddard has research interests in the structure, mechanism and engineering of proteins and enzymatic catalysts for basic research and biomedical applications. His lab conducts wide-ranging research on gene-specific endonucleases, the creation of suicide genes for biotech and medical applications, structural enzymology and protein engineering. He has coauthored over 160 research articles and reviews in these fields since 1990. In 2004, he was one of several recipients of the Newcomb Cleveland Prize from the American Association for the Advancement of Science (AAAS) for published work in the field of Protein Engineering.

Abstract:

De novo protein design offers access to regions of protein fold space that have not been sampled during natural evolution, regions that may contain solutions to novel structural and functional challenges. De novo design can also produce proteins that are highly robust and stable-favorable attributes for subsequent functionalization by rational means. We recently reported the development and experimental validation of a computational method for design of closed tandem repeat protein architectures (a closed repeat architecture is one in which the N- and C-termini are juxtaposed). We have used this method to create a set of alpha-helical solenoid repeat proteins whose bundle handedness is opposite that found in analogous natural proteins. These designed proteins have toroidal architectures with up to 24 repeats, variable thickness and central pore dimensions ranging from 0 to 60 Angstroms in diameter. We are now expanding the topological diversity of the repeat units to access new geometries and functionalizing the designed scaffolds to create reagents for biomedical and industrial applications. A motivating hypothesis behind this work is that designed will prove to be valuable protein scaffolds for protein array and display by virtue of their solubility, stability tunable symmetry, modularity, and self-reinforcing architectures.

Biography:

Erik D Foehr is a Biotechnology Expert with over 15 years of research and development experience, resulting in numerous publications, patents and innovations that propel the advancement of science, medical treatment and patient care. He has received his PhD in Physiology and Biophysics at the University of California, Irvine and Post-Doctoral Fellowship at the Gladstone Institute of Virology and Immunology. He has a strong scientific background, research capabilities and business acumen that drive effective laboratory operations and project management. His work includes contributions in the areas of regulated bioanalysis and medical device characterization. He has conducted numerous developments, validation and testing studies using a variety of analytical approaches. He has proven leadership of bioanalysis research and drug development projects focused on increasing corporate brand, value proposition and impact to the industry.

Abstract:

Protein therapeutics is a dominant and fast growing sector of the $400 billion global pharmaceutical market. There are over 200 marketed protein pharmaceuticals. Major opportunities exist to bring to market the next generation of biopharmaceuticals. In the era of personalized medicine and targeted drugs (immuno-oncology), novel approaches to engineering proteins/peptides are highly desirable. BioTether Sciences develops enzyme, antibody and peptide therapeutics for cancer, infectious diseases, autoimmune diseases, rare diseases and other unmet medical needs. BioTether Sciences uses novel approaches to increase safety, efficacy and targeting of biopharmaceuticals. The technology involves the tethering of proteins with polymer linkers or using high affinity interactions between ligand-receptor and antibody-antigen. Innovator therapeutics can be greatly improved using this approach or novel tethered therapeutics may be developed. Examples and case studies of engineered proteins will be provided. Peglyation or other polymer linker is used to connect proteins and peptides together thereby increasing molecular weight and bulk. This reduces clearance by the kidney and greatly improves drug half-life. The increased vacancy improves receptor binding. Epitope masking improves specificity of targeting to enhance safety. These approaches have been applied to human growth hormone and therapeutic antibodies. For example, human growth hormone may only last for minutes in the circulation without protein engineering. In another application, an antibody is masked by a tethered antigen that improves targeting.

Biography:

Dhanesh Gadre works in the Purification Process Sciences Department at MedImmune LLC, USA. He designs, develops and executes protein purification processes for the development of biopharmaceutical products such as humanized monoclonal antibodies and recombinant proteins. He performs purification and biophysical characterization and formulation of antibodies and protein reagents in response to the needs of several departments using protein biochemistry and chromatography principles and methods. He also has expertise in analytical biochemistry techniques such as HPLC, gel electrophoresis etc. He has Master of Science degree in Chemical Engineering from Syracuse University, USA.

Abstract:

Endotoxins are complex molecules and a significant impurity risk present in the downstream purification processes. Trace amounts of endotoxins can cause immune responses in humans resulting in fever or hypertensive shock. Endotoxins also interfere with the cell based activity assays, impacting the selection of biopharmaceutical drug candidates. For these reasons, it is very important for academic, research and development labs and manufacturing facilities to ensure that protein samples and products are free of endotoxins. During biopharmaceutical production, endotoxins are usually cleared during the downstream purification process. However, if endotoxins interact with a protein of interest through electrostatic or hydrophobic interactions, they can become difficult to remove. Triton X-100 has showed promise in breaking the endotoxins-proteins interactions. However, in some cases Triton X-100 becomes ineffective at breaking these interactions and is therefore unable to remove endotoxins. In this study, we were able to identify a wash condition on chromatography media using a non-ionic detergent octyl-β-D-1-thioglucopyranoside (OTG). This detergent wash can reduce endotoxins from protein solutions to lower levels than Triton X-100 with similar or better protein recovery. Different classes of proteins were bound to different modes of chromatography media and then washed with a variety of detergents. OTG showed the most promising data among these detergents in reducing endotoxin levels with high protein recovery. We examined the mechanism of action to determine why OTG showed better endotoxin clearance ability than Triton X-100. Triton X-100 affects only hydrophobic interactions but OTG can affect hydrophobic as well as electrostatic interactions between proteins and endotoxins. We also showed the impact of the robust OTG on research cell based assays.

Biography:

Kelly Yan has over 17 years of biopharmaceutical experience in the field of protein biochemistry focusing on protein expression, purification and characterization. Her passion has been on difficult-to-produce proteins, from kinases with different phosphorylation status and activities to protein complexes with conformational changes and functions. She has received her MD degree in Shanghai, China before she started her graduate studies in SFSU and research in Charles Craik Lab in UCSF.

Abstract:

Midkine (MDK) belong to a newly evolving family of secreted neurotrophic and developmentally regulated heparin-binding molecules, which also includes pleiotrophin (PTN). Both proteins are involved in promoting neurite outgrowth. Various studies have shown that MDK is involved in cell mitogenesis, transformation, survival, migration and angiogenesis. Moreover, MDK is over-expressed in a number of human cancers; therefore it has become an attractive target for developing cancer therapeutics. Active recombinant human MDK (rhMDK) can be a critical reagent for cell assay development as well as generation of small molecule inhibitors or antibodies for therapeutic drug discovery. MDK is a cysteine-rich 13 kDa protein containing five disulfide bonds (C15-C39, C23-C48, C30-C52, C62-C94 and C72-C104). Expressing and purifying such a protein can be challenging, mainly because the disulfide bonds have to be properly formed to preserve its activity. Traditionally, proteins with disulfide bonds are purified under a denaturing environment and then re-folded. In this study, we expressed (rhMDK) in the Escherichia coli Origami 2 (DE3) strain, which carries a (trxB-/gor522-) double mutation. Soluble rhMDK was expressed at a high level in this strain and protein purification required only one step of heparin affinity chromatography followed by a size exclusion polishing step, without an inclusion body preparation and refolding process. Seven milligrams of rhMDK with high purity were obtained from a 3 L culture. All 10 cysteines were confirmed to be engaged in correct disulfide bond linkages by mass spectrometry analysis. Activity of purified rhMDK was confirmed by a neurite outgrowth assay using rat cerebellar granule cells. Our study demonstrated a very simple and useful technique for producing disulfide bonds-containing bioactive protein with high purity.

Biography:

Soojin Han has completed her PhD from both KIST and Hanyang University in Korea and Post-doctoral studies from University of Alabama and University of Florida in USA. She has been working as a Scientist of the Department of Cell Culture and Fermentation Sciences at MedImmune, a Maryland based biotechnology development enterprise owned by AstraZeneca. She has much experience and great knowledge in mAb and non-mAb production in prokaryotic and eukaryotic expression platforms and published several papers in reputed journals.

Abstract:

Secretion of heterologous proteins into E. coli cell culture medium offers significant advantages for downstream processing over production as inclusion bodies; including cost and time savings and reduction of endotoxin. Signal peptides play an important role in targeting proteins for translocation across the cytoplasmic membrane to the periplasmic space and release into culture medium during the secretion process. Alpha toxinH35L (ATH35L) was selected as an antigen for vaccine development against S. aureus infections. It was successfully secreted into culture medium of E. coli by using bacterial signal peptides linked to the N-terminus of the protein. In order to improve the level of secreted ATH35L, we designed a series of novel signal peptides by swapping individual domains of modifying dsbA and pelB signal peptides and tested them in a fed-batch fermentation process. The data showed that some of the modified signal peptides improved the secretion efficiency of ATH35L compared with E. coli signal peptides from dsbA, pelB and phoA proteins. In particular, one of novel signal peptides improved the yield of secreted ATH35L by 4-fold in the fed-batch fermentation process and at the same time maintained the expected site for signal peptide cleavage. Potentially, these new novel signal peptides can be used to improve the secretion efficiency of other heterologous proteins in E. coli. Furthermore, analysis of the synthetic signal peptide amino acid sequences provides some insight into the sequence features within the signal peptide that influence secretion efficiency.

Biography:

Andrew Lobashevsky MD,PhD, D(ABHI)- Diplomat of American Board of Histocompatibility and Immunogenetics is Associate Professor and Histocompatibility Laboratory Director at Department of Medicine of Indiana University and Clarian Health Partners Inc. Dr. Andrew Lobashevsky joined the IU Department of Medicine in December 2004, as the Directory of the Immunology-Histocompatibility Laboratory. Previously, Dr. Lobashevsky was the Co-director of the HLA Laboratory at the University of Alabama at Birmingham Transplant Center, the third largest center in the country. Dr. Lobashevsky received his medical education and postgraduate training at the Department of Immunology and Microbiology of Sechenov’s Medical Academy, Moscow, Russia. He had his post-doctoral training in cellular and molecular immunology at the University of Tennessee at Memphis, and received transplant immunology/histocompatibility training at the University of Alabama at Birmingham. The laboratory headed by Dr. Lobashevsky is focused on providing service for the solid organ transplant programs as well as the bone marrow transplant activities of the Hematology/Oncology division at IU.

Abstract:

In solid organ transplantations the graft outcomes critically depend on the degree of human leukocyte antigen (HLA) matching between the donor and recipient. Although the cellular component of the immune response to the transplanted tissue plays a key role, the contribution of antibodies should not be underestimated. Highly sensitive technologies such as solid-phase based single antigen assay allow to determine even low concentrations of donor specific antibodies and with a high degree of confidence to predict graft outcomes. Development of anti-HLA antibodies strictly depends on immunogenicity of mismatched HLAs. The latter is defined by fi ne epitope structure of HLA. Each HLA protein represents a linear sequence of amino acid residues (AAR) or triplets and the degree of mismatch is assessed as the number of triplets that are not shared between the donor and the recipient. Further investigations of the three dimensional structure of antibody-antigen complexes showed that HLA epitopes could be presented by a group of AARs that are not located beside one another, but rather represent a 3-Å to 5-Å radius patch. These patches have been defined as eplets. Some of eplets include short sequences of AARs, which are equivalent to triplets, whereas, others contain discontinuously located AAR. Further studies demonstrated that area of interaction between complementarity determining region of antibody and HLA is about 900 Å2 and comprises structural and functional epitopes. The former is responsible for binding, whereas the later determine strength of antigen-antibody interaction, which intern results to conformation changes of antibody and subsequent complement activation.

Biography:

Itai Benhar is a full Professor, Vice Dean for Research at the George S Wise Faculty of Life Sciences, Tel-Aviv University, Israel. He received in 1992 a PhD in Molecular Biology from the Hebrew University, Israel and was a Post-doc 1992-1995 at NCI, NIH. He joined Tel-Aviv University as a tenure-track assistant professor in 1995 and is a full professor since 2008. He is an expert in the fields of antibody engineering, drug discovery and drug delivery with over the 20 years of being active in these fields, 92 peer reviewed articles and 11 patents.

Abstract:

Small interfering (si) RNAs can be used to silence disease-causing genes. However, their development as drugs has been limited mainly to knocking down liver gene expression, since delivery to other tissues requires development of a targeted delivery carrier. Modulating immune cells function using siRNAs holds great promise in advancing targeted therapies to many immune-related disorders including cancer, inflammation, autoimmunity, and viral infections. However, the ability to effectively knockdown gene expressing in leukocytes is still challenging. Here we present a modular platform to target specific cell types, exemplified here with immune cells, using siRNA loaded lipid nanoparticles (LNP) coated with oriented, targeting antibodies non covalently bound to a membrane-anchored lipoprotein that recognizes their Fc domain. Unlike chemically conjugated antibodies, these oriented antibodies maintain their high affinity and the LNPs avoid scavenging by Fc receptors on macrophages. A simple switch in 5 different targeting antibodies (against Ly6C, CD3, CD4, CD25 and Itgb7) redirected the LNP for exquisitely specific uptake in diverse leukocyte subsets in vivo and enabled specific knockdown in difficult-to-transfect CD4+ cells. Intravenously injected anti-Ly6C-coated LNP encapsulating TNF siRNAs were taken up selectively by Ly6C+ monocytes and activated tissue macrophages, suppressed TNF-alpha expression in the colon and ameliorated inflammatory bowel disease symptoms in a DSS-induced colitis mouse model, demonstrating the platform’s potential therapeutic utility. This approach opens new avenues for studying cell biology in vivo and potentially for a wide range of therapeutic applications in a cell-specific manner.

Biography:

Anna V Hine has studied at the University of Manchester (UK) and Harvard Medical School. She is a Reader and Associate Dean Enterprise at Aston University (UK). In March 2013, she was named BBSRC Commercial Innovator of the Year 2013, for her work in transferring ‘ProxiMAX randomisation’ into SME Isogenica Ltd. A Molecular Biologist by training, she relishes interdisciplinary work.

Abstract:

ProxiMAX’ randomization is the technology that lies behind Isogenica’s Colibra™ offering. It is a defined saturation mutagenesis process that delivers precision control of both identity and relative ratio of amino acids at specified locations within a protein/antibody library. Thus unwanted amino acids such as cysteine and methionine can be eliminated from libraries because no constraints are imposed by the genetic code. Moreover, the process is non-degenerate, which means that encoding DNA libraries are as small as is physically possible. ProxiMAX relies on a process of saturation cycling comprising ligation, amplification and digestion for each cycle and is the science behind the commercial “Colibra™” technology. With achieved diversities of >99% (6 and 11 saturated codons) and the potential to generate libraries of up to 1014 components, we contest that ProxiMAX randomization is a vital tool in engineering antibody libraries of the highest quality. Th is presentation will examine the development of the ProxiMAX process and give examples of antibody libraries created to date.

Biography:

Ernesto J Fuentes began his Post-graduate education at the University of Dayton (Dayton, Ohio) with a Master's degree in Developmental Biology. He has obtained a PhD degree in Biochemistry from University of Illinois under the mentorship of Dr. A Joshua Wand. He has pursed Post-doctoral training at the University of North Carolina in the areas of protein NMR dynamics with Dr. Andrew Lee and Rho-family GTPase function with Dr. Channing Der. His current research broadly focuses on elucidating the molecular mechanisms that regulate signal transduction. Our recent work has focused on two systems: the regulation of Rho-family GTPase signal transduction and bacterial two component systems.

Abstract:

Molecular recognition is critical for the function and regulation of signal transduction in all cell types. PSD-95/Dlg1/ZO-1 (PDZ) domains are among the most abundant protein-protein interaction modules in the human proteome, commonly found in multi-domain signal scaffolding proteins. PDZ domains typically recognize a variety of short amino acid motifs, including the C-termini and internal peptide sequences of partner proteins. How PDZ domains accommodate these diverse interaction partners while providing specificity remains poorly understood. The overall goal of our studies is to define the molecular basis underlying PDZ domain specificity towards its known ligands. In this presentation, I will discuss recent studies that reveal, how specificity is obtained and rationally altered in model PDZ domains that bind C-terminal and internal peptide sequences? In addition, I will provide examples for how conformational dynamics and structure both contribute to molecular recognition of altered PDZ proteins? These studies have important implications for the evolution, design and regulation of protein-ligand interactions.

Biography:

James Endrizzi, founder of SuperNova C, is a Ph.D. chemist and X-ray crystallographer based in Missoula, Montana. He specializes in the use X-ray crystallography combined with biochemical, biophysical, and bioinformatic approaches to unveil structure-based molecular mechanisms for macromolecular function. With a B.S. in Chemistry & Biology from the University of Minnesota and a doctorate from Eugene, Oregon, James went on to a post-doctoral position in the Alber lab at UCBerkeley, where the newly commissioned tunable, synchrotron beam 8.3.1 was available at the Atomic Light Source for many Friday and Saturday nights spent experimenting on various crystals. After 9 years at Berkeley, he went on to research stints at UC Davis (4 years),, Montana State University (3 ½ years) and the Hormel Institute, giving 15 talks, presenting over 20 posters and publishing 23 papers along the way before founding SuperNova C, a company devoted to providing vitamin C as mineral ascorbates.

Abstract:

Aspartate transcarbamoylase (ATCase) has been featured in many biochemistry textbooks as an example of allosteric enzyme regulation. It catalyzes the fi rst step of pyrimidine nucleotide biosynthesis and is feedback inhibited by cytidine triphosphate (CTP), which is the end product of the pathway. ATCase exhibits cooperative substrate binding to its catalytic subunits and allosteric binding of nucleotides to its regulatory subunits. Opposing inhibition by CTP and activation by adenosine triphosphate (ATP) promote homeostasis of purine and pyrimidine nucleotides. Regulation of ATCase activity has been interpreted as a ligand promoted change in the equilibrium between inactive (taut) and active (relaxed) states largely consistent with a two-state allosteric model. The structure of the ligand-free enzyme represents the inactive, taut state and the structure of ATCase bound to a bisubstrate analog (N-phosponacetyl-L-aspartate; PALA) has been proposed to represent the active, relaxed state. However, there is little evidence that the PALA-bound enzyme is structurally similar to the active conformation(s) in the absence of substrates. A novel approach to define the structure of the ligand free relaxed, or “activated” state is to use mutant enzymes that destabilize the taut state and shift the allosteric equilibrium toward the relaxed-state conformation(s). In contrast to the wild-type enzyme, which exhibits a more compact global conformation and sigmoidal enzyme kinetics, the mutants are more expanded and display hyperbolic kinetics characteristic of non-cooperative enzymes. Thus, we tested the hypothesis that ATCase is activated through the modulation of flexibility by determining the X-ray crystal structures of several activated-state mutants. We obtained crystals of three ATCase mutants in multiple crystal forms, collected X-ray diffraction data on the various forms and solved and refined dozens thus far. A variety of tertiary and quaternary structures in activated-state mutant enzymes support the hypothesis that these structures represent metastable states accessed by an ensemble of activated-state conformations..

Biography:

Seth McDonald received his PhD in Biochemistry from the University of Utah where he studied the structure and biological implications of the interaction of the essential Spn1 and Spt6 transcription factors in Saccharomyces cerevisiae. In his Post-doc at Colorado State University, continuing his use of x-ray crystallography and enzyme kinetics, he engineered fast and accurate RNA polymerases from small RNA viruses that effectively restrict viral genome variations within the given population required for infection and survival. At Illumina, he has made significant contributions to the structural and functional understanding of how polymerases incorporate nucleotides modified on the base and sugar. He is the assay development and structural biology lead in the protein engineering group where he continues to contribute to the ongoing enzyme engineering projects within Illumina.

Abstract:

The sequencing-by-synthesis (SBS) chemistry commercialized by Illumina has been a key enabler of massively parallel nextgeneration sequencing, which in turn has dramatically reduced the cost of sequencing human genomes and resulted in personalized medicine initiatives and population sequencing efforts throughout much of the world. The capacity of engineered archaeal family B DNA polymerases (pols) to incorporate modifi ed nucleotides has been exploited by researchers throughout academia and biotechnology, including Illumina’s SBS chemistry. Nucleotides with a 3’-reversible terminator and fluorophores attached via the base serve as the basis for base calling in Illumina NGS platforms. Pols deployed on the fi rst Genome Analyzer™ instruments had poor incorporation effi ciency of these modifi ed nucleotides. Through our engineering eff orts, we have facilitated a significant reduction in chemistry time while improving sequencing data quality and enabling over 300 bp read lengths. The critical goal of our research is to engineer pols with enhanced selectivity, fidelity, and incorporation efficiency of modified nucleotides to enable faster sequencing turnaround time and boost adoption of NGS technologies in the clinic.

Biography:

Xiaoying Zhang is Professor at Northwest Agricultural and Forestry University (elite university in the national 985 and 211 programs), China since 2008. He is graduated from China Pharmaceutical University (undergraduate study in Pharmacology) and Charité Medical School (Post-graduate and PhD in Pharmacology) of Freie Universität Berlin and Humboldt-Universität Zu Berlin, Germany. Before and during his PhD study, he has worked in WHO- headquarters Geneva and Robert- Koch Institute Berlin. He was the nominee for the outstanding Young Scientist Award, China (2015). His major research interests are: antibody engineering, immunoassay and rapid detection/diagnosis, nature product research and pharmacological study. He has contributed 100+ publications (from which 80+ international publications). He is holding three Chinese patents for his innovative research accomplishments. He has also edited fi ve books (four– Chinese and one in English) and contributed three book chapters in the books of International publishers. He has been appointed as editor and/or reviewer in more than 40 international journals and serves as Member of Board of Examiner in universities from China, Germany, India and Pakistan.

Abstract:

Residues of antibiotics in the aquatic environment and in food of animal origin represent a major concern, as prolonged exposure to antibiotics is a serious health hazard, related to both the side effects of prolonged use and the risk of developing bacterial resistance to various antibiotics. Given the low levels of the antibiotics residues in complex matrices, the development of sensitive analytical methods represents a major challenge. We report the current state of the art in the determination of antibiotics based antibody engineering and immunoassays. First, diversified antibody generation platforms/methods promote the screening and assessment of high qualified antibodies. Second, various immunoassays simplify and empower the immune-detection applications for different biomedical purposes. Third, there are tremendous academic motivations and industrial needs in the detection/evaluation of antibiotics by using antibodies and immunoassays. There are diversified antibody generation platforms/methods (e.g., Polyclonal antibody (pAb), Cell engineering antibodies—hybridoma technology, recombinant antibdy (rAb)). Different species of antibodies e.g. chicken antibody (polyclonal IgY, monoclonal IgY), camel antibodies, and rabbit antibodies have been widely used. The recent advancements in antibody engineering permit to produce chicken mIgY quickly and inexpensively by phage display technology, because chicken possess only one functional immunoglobulin heavy chain variable region (VH) gene and one light chain variable region (VL) gene. The purpose of our study is to prove that the mIgY could be used for immunological detection and diagnosis, for screening and validating of biomarkers and drug targets. There is a huge demand for the detection reagent of antibiotic residues on the market, and it gradually becomes the intrinsic driving force of the development of industry. Diversified antibody generation platforms/methods promote the screening and assessment of high qualified antibodies. Various immunoassays simplify and empower the immune-detection applications for different biomedical purposes.

Biography:

Amirali Kia has completed his Bachelor’s and Master’s working on the application of various artifi cial intelligence algorithms in computational fl uid mechanics. He then started his PhD at Stanford University where his focus was on high performance computing and developing fast algorithms with applications in computational biology. He started his career at Illumina by joining Protein Engineering group to apply machine learning algorithms to enzyme engineering. He is now a senior manager at Illumina in the Computational and Applied Biology Department, leading Deep Learning group. The focus of his team is to bring new AI algorithms to genomics and Illumina’s internal technology.

Abstract:

Statement of the Problem: Next generation sequencing has opened new doors into genomics. Recent advances in sequencing technology have enabled researchers to sequence genomes with unprecedented accuracy and speed. However, preparing DNA sequencing libraries still remains as a challenging step. Transposase-based library preparation off ers a simple, fast, scalable, and flexible solution. However, when compared to other (ligation-based) library preparation methods, transposases show some insertion bias that aff ects the fi nal DNA sequencing library.

Methodology: We applied various machine learning algorithms to random mutagenesis libraries of transposases to efficiently design new mutant transposases with less insertion bias.

Findings: We present the discovery of a mutant transposase (Tn5-059) with low GC insertion bias and AT dropout. Libraries prepared by this mutant have good uniformity of genome coverage as well. Tn5-059 also shows low sensitivity to the amount of input DNA. In addition, this enzyme shows effi cient performance in DNA sequencing libraries for open chromatin profiling.

Conclusion & Significance: DNA input tolerance together with superior coverage uniformity are two important factors to be considered in DNA sequencing library preparation. Tn5-059 successfully delivers on both of these aspects, which leads to less sequencing volume and lower sequencing cost. We discuss the importance of choosing a correct assay as well as the importance of filtering data based on the biology behind the assay.

Biography:

Steven Alexander Mann received his Medical degree in 2015 from the University of Alabama, School of Medicine and is currently a Pathology Resident at the Indiana University School of Medicine. He is actively involved in managing molecular requests for his current institution and is a Member of the College of American Pathologists Engaged Leaders Network. He has multiple peer-reviewed publications on a variety of topics including a recent review of PD-L1 immunohistochemistry for genitourinary tumors and a chapter on biomarkers for hepatic and pancreatobiliary malignancies

Abstract:

Novel immunotherapies have recently demonstrated significant objective responses in many advanced malignancies. The standard therapy of some of these cancers had remained unchanged for decades despite poor outcomes. Immune checkpoint inhibitors, in particular those for the programmed cell death protein (PD-1) pathway, have bought great progress for the care of these patients in the past decade. Currently, the most clinically significant ligand of PD-1 is the programmed cell death ligand-1 (PD-L1). When this pair binds, cytotoxic T cell function is inhibited. Many types of tumors can express PD-L1 as a mechanism to evade the antitumor response of the immune system. By the end of 2016, the United States Food and Drug Administration (FDA) had approved four different immune checkpoint inhibitors for use in a total of six different types of malignancies. Many of these drug-indication combinations have an FDA-approved companion PD-L1 immunohistochemistry assay available. These PD-L1 assays are used to predict therapeutic responses and in some cases tumor PD-L1 positivity is a prerequisite for initiating therapy. Each of these assays uses a different anti-PD-L1 clone. The use of these antibodies and the interpretation of tumor PD-L1 expression are continually evolving, as well as the understanding of the overall impact of this information. Knowledge of the current state of these rapidly evolving drugs and assays will prepare researchers, manufacturers and clinicians for future applications of these promising antibodies.

Biography:

Estefania Arevalo-Tristancho is currently pursuing Master’s degree in Design and Process Management with emphasis on Bioprocesses.

Abstract:

Enzymes of microbial origin have been proven to be useful in different fields such as medicine and the food industry. L-asparaginase, of microbial origin is an amidohydrolase enzyme, which catalyzes the conversion of L-asparagine to aspartate and ammonium cation. L-asparaginase is known as an anti-cancer agent, which prevents the proliferation of tumor cells by decreasing the level of asparagine in the blood. Th is enzyme has been shown to be a form of treatment for acute lymphocytic leukemia (ALL), extracted from E. coli and Erwinia chrysanthemi, which have a high commercial value and multiple side effects due to L-glutaminase activity produced by them. In this research, 25 Streptomyces isolated from the Arauca River bank (Equatorial zone, Colombia) with L-asparaginase activity were found by the Nessler method. Studies have shown that Streptomyces, in addition of being a source of easy access and production, produce L-asparaginase with less or none L-glutaminase activity. Streptomyces isolated were identified morphologically and molecularly. Plackett-Burman design established that at 30 ºC, 200 rpm, pH 7, lactose (1%) and malt extract/asparagine (0.15%) were the best conditions for the fermentation of the isolates and enzyme production. This enzyme secreted in the medium will be purifyed by dialysis, lyophilization and ion exchange chromatography to establish a mathematical model to simulate the effect of the substrate, pH and temperature on L-asparaginase activity.

Biography:

Marissa L Matsumoto is a Scientist at Genentech, Inc., in the Department of Structural Biology. Her lab focuses on Protein and Antibody Engineering to develop novel research tools for the study of complex ubiquitination events in cell signaling. She has obtained her PhD from Washington University in St. Louis and her BA from Northwestern University.

Abstract:

Statement of the Problem: Ubiquitin is a post-translational modification involved in nearly every signaling pathway. Monoubiquitination occurs when the carboxy-terminus of ubiquitin is linked through an isopeptide bond to a lysine residue on a substrate. Ubiquitin itself contains seven lysine and a free amino-terminus through which additional ubiquitin subunits can be linked, resulting in polyubiquitin chains of different topologies. Determination of polyubiquitin chain linkages requires the use of ubiquitin mutants or complex mass spectrometry experiments. The purpose of this study is to engineer antibodies to detect specific polyubiquitin linkages to provide useful research reagents for the study of ubiquitination in cells.

Methodology & Theoretical Orientation: We used phage display to engineer antibodies with exquisite specificity to the linear, K11, K48, and K63 linkages and used X-ray crystallography to elucidate the nature of their specificity. To detect more complex heterotypic chains containing mixed or branched linkages, we have developed bispecific antibodies using the knobs-into-holes technology.

Findings: The antibodies are highly specific for a given linkage and work in numerous applications, including western blot, immunoprecipitation, and immunofluorescence. Epitope determination by X-ray crystallography demonstrates that rather than contacting the linkage itself, the antibodies recognize a conformational epitope that results from the relative orientation of two ubiquitin subunits because of the spatial positioning of the residue involved in the linkage.

Conclusion & Significance: The polyubiquitin linkage-specific antibodies have provided ubiquitin researchers with an easy-to-use reagent to quickly and efficiently determine the linkages of polyubiquitin chains without the need for ubiquitin mutants or access to mass spectrometry equipment and expertise. These antibodies have become essential tools in studying ubiquitination in cell signaling and have aided in elucidation of numerous pathways including polyubiquitin chain editing, K11-linked chains in cell cycle control, and K11/K48-branched chains in cell cycle and protein quality control.

Biography:

Stephen D Kirby has been working in the field of chemical threats for the US Army Medical Research Institute of Chemical Defense, Aberdeen Proving Grounds for 30 years, to include areas of research such as cyanide detoxification, sulfur mustard metabolism, and edemagenic pulmonary threats. The focus of his current research area has been developing and evaluating catalytic and stoichiometric protein bioscavengers for organophosphorus compounds. Current research has investigated enzyme platforms that include human carboxylesterase, human paraoxonase-1, platelet activating factor acetylhydrolase and the bacterial enzyme phosphotriesterase.

Abstract:

Organophosphorus compounds (OPs) such as sarin and soman are some of the most toxic chemicals synthesized by man. They exert toxic effects by inactivating acetylcholinesterase (AChE) and bind secondary target protein. Enzymes can be engineered by amino acid substitution into OP-hydrolyzing variants (bioscavengers) and used as therapeutics. Some enzymes associated with lipoproteins, such as human plasma platelet-activating factor acetylhydrolase (pPAF-AH), are also inhibited by OPs; these proteins have largely been ignored for engineering purposes, because of complex interfacial kinetics and a lack of structural data. We have expressed active human pPAF-AH in bacteria and previously solved the crystal structure of this enzyme with OP adducts. Using these structures as a guide, we created histidine mutations near the active site serine of pPAF-AH (F322H, W298H, L153H) to generate novel OP-hydrolase activity. Wild-type pPAF-AH, L153H, and F322H have essentially no hydrolytic activity against the nerve agents tested. In contrast, the W298H mutant displayed novel somanase activity with a kcat of 5 min-1 and a KM of 590 μM at pH 7.5. There was no selective preference for hydrolysis of any of the four soman stereoisomers. The kcat/KM for W298H is 8x103 M-1 min-1, a significant enhancement over the wild-type enzyme

Biography:

Karen Bunting is a Science Director at Albumedix, heading Molecular Biology and Fermentation within Research and Technical Development. Prior to the formation of Albumedix, she has joined Novozymes Biopharma UK in 2011 as a Senior Research Scientist and later as Science Manager. She has 20 years’ experience in Structural Biology and Protein Engineering. At Albumedix her primary research focus has been the design and engineering of albumin variants to optimize the circulatory half-life of therapeutic agents. Prior to joining Novozymes, she led a research team at the University of Nottingham focusing on structural analysis of protein-protein and protein-DNA interactions, following on from Post-doctoral work at the Institute of Cancer Research in London. She has obtained BSc in Microbiology from Imperial College, London and PhD in Crystallography from Birkbeck College, London.

Abstract:

Human serum albumin is the most abundant serum protein and has a long history of safe use in biopharmaceutical products. It has many features that contribute to the molecule being naturally well designed as a drug carrier, including its high solubility and stability, long circulatory half-life and the potential to attach therapeutic candidates by fusion or conjugation. We will describe Veltis® albumin variants designed to enhance two of these aspects, long circulatory half-life and drug attachment by conjugation, whilst maintaining solubility, stability and its inherent low immunogenicity. The long circulatory half-life of albumin in the human body (around 19 days) derives in part from the size of the molecule, whereby it resists filtration through the kidneys and in part from its association with the neonatal Fc receptor (FcRn). Albumin binds to FcRn under acidic conditions in the endosome and is rescued from lysosomal degradation back to the plasma where it is released under neutral pH conditions. We will describe our rational design of variants to enhance binding to the receptor under acidic conditions, whilst retaining release at neutral pH, resulting in a greater than two-fold improvement in half-life. Short half-life is a significant challenge for many peptide and protein therapeutics; fusion or conjugation to Veltis engineered albumins offers the potential of monthly dosing. We will also present thio-engineered albumin variants to improve drug efficacy by increasing the drug-albumin ratio whilst maintaining FcRn-binding capabilities and half-life extension. Rational protein design has enabled us to enhance specific properties of albumin related to drug delivery whilst retaining the numerous other features which contribute to the molecule being naturally well designed as a drug carrier.

Biography:

Christopher H Gray has obtained his BSc and PhD from the University of Glasgow, examining the biochemical basis of multidrug resistance in pathogenic fungi. He has then moved to the Institute of Cancer Research in London to gain experience as an X-ray Crystallographer where he was trained in a diverse range of protein production techniques. Following this he took up a position at the CRUK Beatson Institute becoming an early Member of Staff in the Institute’s Drug Discovery Program. As a Structural Biology Team Leader he has day to day responsibility for the protein production, NMR, biophysics and X-ray Crystallography in the program.

Abstract:

The protein production section of the Beatson Institute’s Drug Discovery Program supplies a considerable number of highly purified and active recombinant proteins for structural biology, biophysical and biochemical applications. Pressure to satisfy high demand has prompted the development of novel and innovative methods to streamline workflow maximize output and ensure rapid delivery of critical proteins. Driving up soluble yields, ensuring productive cell culture and reducing the burden on purification resources can enhance the output of a protein production facility. We have substantially improved our productivity by implementing a series of new bacterial MBP fusion vectors. These systems substantially elevate soluble expression but also co-express a protease that removes the MBP tag in vivo, affording the benefits of MBP fusion on soluble yield but eliminating the MBP moiety from downstream purification. As an extension of this, we have developed several systems that allow the monitoring of recombinant expression during fermentation, using an auto-cleaved fluorescent protein tracer. This allows the operator to infer adequate target expression in near real-time, ensuring that cultures allowed occupy fermentation resource lead to satisfactory product yields. We have also accelerated productivity by reducing the number of chromatography steps in common protocols and automating FPLC purification. Taking our inspiration from several emerging and newer multimodal chromatography technologies, we have developed rapid, mixed chemistry affinity strategies to purify tandem tagged proteins. This allows the isolation of high purity target in a single column, often removing the need for slow size exclusion chromatography to polish the product. Finally we have implemented routine multidimensional chromatography on our AKTA AVANT systems performing complex sequences of protein purification and conditioning steps with a minimal requirement for user intervention. The net result is more reliable and frequently higher yield preps, delivered to the downstream user in the shortest possible time.

Biography:

Haiyan has over 19 years of industry experience in the pharmaceutical and diagnostics areas. Haiyan joined Centocor R&D in 2004, which was later named Janssen BioTherapeutics.  In the past 13 years at Janssen Haiyan leads a protein biochemistry group, focusing on developing and optimizing biotherapeutic lead molecules.  She leads the group to work on purification and characterization of monoclonal antibodies, alternative scaffolds including bispecific antibodies, target antigens, receptors and other proteins for various research projects; developing new technologies to improve HTP purification and biochemical QC analyses; aligning with the methods from Discovery to Development process to ensure the smooth transfer of the therapeutic candidates from Lead optimization to Development stages. Haiyan received her Ph.D. in Microbiology at the University of Iowa.  She then took a postdoctoral position at Eli Lilly and Company (Indianapolis).  From 2000 to 2004, Haiyan worked at Roche Diagnostics Division (New Jersey) to lead a group responsible for developing purification process and manufacturing of recombinant DNA polymerases which were used for disease diagnosis.

Abstract:

Monoclonal antibody therapies continue to be the focus of biopharmaceutical industry.  Hundreds of antibodies are in pre-clinical and clinical development. Purification and characterization of the lead molecules is vital during the lead optimization and candidate selection stage.  Protein A affinity chromatography and additional polishing steps are used to purify panels of antibody hits against specific target.  Throughout the process, biochemical and biophysical assessment for developability and manufacturability are applied to select lead candidate.  This talk will review the methods, the challenges, and how the purification and characterization data contributes to lead selection.  A case study will be discussed on a small scale purification method scouting for difficult proteins which facilitated the development of the large scale process and shortened development timeline.

Biography:

Jeff Mihailoff has an MS in Molecular Biology and MBA from the University of Florida. Since leaving academia, he has been selling research tools into the Biopharma and Academic research market. Prior to ForteBio, he was the bioprocess and chromatography specialist at GE Healthcare focused on downstream purification. He also has experience with multiple different bioanalytical techniques such as Surface Plasmon Resonance, Isothermal Titration Calorimetry, and HPLC. He and his wife enjoy scientific advancement and have dedicated their lives to assisting scientists in taking their technology to the next level.

Abstract:

Pall Fortebio’s Bio-Layer Interferometry has been utilized by the antibody biopharma market since its inception in 2005. This game-changing technology allowed scientists to bypass ELISA and HPLC technologies for antibody quantification. In addition, its lower price point and easy of use allowed it to be a substitute for Surface Plasmon Resonance (SPR) studies on antibody-antigen binding affinity. However, there have remained questions about the accuracy of the work as related to SPR. In this talk, the author will review a recent large-scale study in which different variables related to assay design were tested and then compared to SPR studies. Finally, the author will review some of the newer technology changes and how they are being applied to bioprocess residuals quantification.

Biography:

Thomas Rohrer is Associate Director of Bioconjugate Commercial Development at Lonza and has over 30 years of experience in biotherapeutic process development, scale-up and manufacturing. He holds a BS in Biochemistry and MS in Chemical Engineering. He established process development and clinical manufacturing of ADC(s) at Cambrex Biopharma in 2005. After Cambrex Biopharma was acquired by Lonza in 2007 he joined the Lonza Exclusive Synthesis ADC business team in Visp, Switzerland. Prior to joining Lonza he held senior positions in biotherapeutics process development and manufacturing at Human Genome Sciences, Otsuka Pharmaceutical and the National Cancer Institute (NCI) - Frederick Cancer Research Facility. 

Abstract:

Advances in the coupling of antibodies to potent cytotoxic drugs have resulted in stable delivery platforms with improved pharmacokinetics, which spares patients from the debilitating systemic toxicity observed with traditional chemotherapy. Two ADCs Adcetris® (brentuximab vedotin) and Kadcyla (trastuzumab emtansine) are currently approved for difficult -to-treat therapeutic indications. Currently three more ADCs, Inotuzumab ozogamicin (Pfizer),Vadastuximab talirane (Seattle Genetics) and Depatuxizumab mafodotin (AbbVie) are undergoing regulatory review or completing Phase III clinical studies. In all, more than 60 ADCs are in clinical trails with over 100 more in the pipeline. In addition to ADCs the multi-faceted field of bioconjugates also extends to coupling targeting agents to non-cytotoxic payloads. Perserving the targeting ability of antibody and the effector function of the payloads require that process R&D find the intersection of reaction parameters which preserve the integrity of both molities. Case studies will be presented which discuss the development and manufacturing of several next generation bioconjugates. 

Biography:

Victor Guzov has his expertise in protein engineering and evolution, mode of action of insecticidal proteins and genetic engineering of crop plants. He has been long involved in optimization of insecticidal proteins for the purpose of protection of crop plants from insect infestation using biotechnology approaches. That led him to research on mode of action of insecticidal proteins, and ways to assess their value for insect resistance management. His collaborative research with both industry and academic scientists on evolution of Bt proteins resulted in new opportunities for effective management of insect resistance to genetically-modified crop plants.

Abstract:

The development of insect resistance to insecticidal proteins, such as Bt proteins, is a continuous threat to insect control traits in biotech agricultural crops. This requires constant search for insecticidal proteins with new modes of action (MOA), which enhance insect resistance management options via delaying new resistance development and combating already existing resistant insects. This presentation will describe the successful onboarding of phage-assisted continuous evolution (PACE) technology, and its application to Bt toxins for generating new MOAs.

Biography:

Yvette Stallwood has completed her PhD at the University of Birmingham (UK) and joined Lonza in 2007. She has been Head of Applied Protein Services for five years. The Applied Protein Services team are focussed on the development and provision of services to support the development of new biotherapeutic proteins with a particular focus on immunogenicity and manufacturability.

Abstract:

This presentation will discuss how in silico and in vitro methodologies are employed to perform a developability and immunogenicity risk assessment in order to highlight potential risks of failure for the development of biotherapeutics. In silico methods can be used to evaluate protein sequence and structure to assess the likelihood of immunogenic responses and potential manufacturability issues including aggregation and PTMs. Ex vivo T and B-Cell responses enable the assessment of overall immunogenicity risks; different approaches are highlighted to further identify processed and presented epitopes.

Biography:

Franz Grus is the Head of Experimental Ophthalmology, Department of Ophthalmology in Johannes Gutenberg University Medical Center, Germany. His area of interest includes: Glaucoma, age-related macular degeneration and dry eye biomarker research, high-throughput-research methods, proteomics and immunology, diagnostic technologies, antibodies in diagnosis and treatment in personalized medicine. He has been in positions like: Principal investigator for several research projects funded by the German Research Foundation (DFG) and the German Society of Ophthalmology (DOG), as well as other institutions and companies.

Abstract:

In glaucoma, the elevated intraocular pressure cannot explain the disease in all patients. Glaucoma is a neurodegenerative disease, leading to the loss of retinal ganglion cells (RGC). Immunoproteomics could play a significant role. Several studies could provide hints for an involvement of autoantibodies in the pathogenesis of the disease. One of those candidates is several heat-shock proteins AAB. The complex profiles of antibodies were analyzed by mass spectrometry based techniques and customized antigen microarrays of 40 different antigens in more than 1000 patients. The resulting profiles were analyzed by different data mining techniques such as artificial neural networks. In all studies we could demonstrate consistent up- and down-regulations of immune reactivities against ocular antigens in glaucoma. The glaucoma could be recognized by a sensitivity and specificity of more than 90%. Furthermore, these antibodies could be useful as an innovative glaucoma treatment option. We analyzed in a glaucoma animal model the effect of an intravitreal injection of anti-synuclein antibodies. Axon density/mm² showed a decay (p<0.01) in controls, (p<0.01) in buffer group, and (p=0.19) in the α-synuclein group. Mass spectrometry revealed changed levels of CCDP93 (-2.6x), cofilin-1 and reticulon 4 (both -2.5x) in IOP elevated eyes and peripherin-2 (2.4x), cofilin-1 (3.5x) and malate dehydrogenase (11.9x) in α-synuclein treated eyes. The results of this study demonstrate clearly that antibody patterns could be useful for diagnosis especially if transferred to a point-of-care device such as lateral flow assays, but also given intravitreal, a promising new approach for neuroregenerative treatment in personalized medicine in glaucoma.

Biography:

Anna V Hine has studied at the University of Manchester (UK) and Harvard Medical School. She is a Reader and Associate Dean Enterprise at Aston University (UK). In March 2013, she was named BBSRC Commercial Innovator of the Year 2013, for her work in transferring ‘ProxiMAX randomisation’ into SME Isogenica Ltd. She is a Molecular Biologist by training.

Abstract:

ProxiMAX and MAX randomization technologies are defined saturation mutagenesis processes that deliver precision control of both identity and relative ratio of amino acids at specified locations within a protein library. Both processes are non-degenerate, meaning that encoding DNA libraries are as small as is physically possible. ProxiMAX is the technology that lies behind Isogenica’s Colibra™ offering and is ideal for saturating contiguous codons, as required in antibody libraries. In contrast, ‘MAX’ randomization targets codons at disparate locations within a gene and is therefore more applicable to other scaffolds or proteins. Since no constraints are imposed by the genetic code, both technologies can eliminate unwanted amino acids such as cysteine and methionine from libraries or encode desired subsets of amino acids with ease. Yet their underlying processes are quite different. This presentation will examine the development of both ProxiMAX and MAX randomization process and give examples of libraries created to date.

Biography:

Stefan Zielonka has received his PhD from the Technische Universitaet of Darmstadt, Germany, where he worked in the Group of Harald Kolmar in the field of Protein Engineering of non-canonical antibodies. Now, he works as Senior Scientist at Merck KGaA (EMD Serono), Germany, in the Department Protein Engineering and Antibody Technologies (PEAT). He was Fellow of the Merck’sche Gesellschaft fuer Kunst und Wissenschaft e.V. and was awarded with the Kurt-Ruths-Award (2016) and the Rainer-Rudolph-Award in Biotechnology and Protein Chemistry (2016).

Abstract:

Yeast surface display has proven to be a versatile platform technology for antibody engineering enabling online and real-time analysis as well as characterization of library candidates. During this talk, the process of antibody library generation will be explained as well as selection of target-specific antibodies with prescribed properties since desirable features such as species cross-reactivity can be implemented into the screening procedure using FACS. Additionally, a novel streamlined one-step approach for the generation of yeast surface display Fab libraries will be discussed that allows for simultaneous introduction of heavy chain and light chain variable regions into one single display vector. Finally, a generic approach for the generation of human IgG-like bispecific antibodies will be presented that relies on the combination of immunization of transgenic rats with yeast surface display.

Biography:

Lia Monica Junie is the Head of the Department, also coordinating the activities of both Laboratory Medicine specialty resident doctors and PhD doctor’s thesis in the medicine field. She is a Member in the Board of Scientific Societies, Reviewer in many peer-reviewed journals, Expert of the Ministry of Education and Research and Evaluator. She has coordinated research projects, published books and more than 200 scientific articles in prestigious journals. She has organized and attended  numerous national, international congresses, as President, Member in the Organizing Committees, Invited speaker, Keynote speaker or Chairperson

Abstract:

Background: Staphylococcus aureus infections remain a serious medical problem and prompt initiation of appropriate antimicrobial chemotherapy is important to improve the prognosis of these infections.

Methods: The strains were isolated from hospitalized patients in Universities Hospitals, Cluj Napoca, Romania, during 2016. Species identification was achieved by standard procedures and by the automated system Vitek 2 (Biomérieux). The susceptibility to antibiotics was performed using the disk diffusion method (Kirby-Bauer) according to the CLSI criteria and by the Vitek 2 System, for a total of 855 strains.

Results: From all isolated strains, 51 (5.9%) yielded Staphylococcus aureus. 34.3% of the S. aureus isolated strains were MRSA. 47.1% of the S. aureus tested strains were resistant to clindamycin. Moderately increased percentages of resistance were found by testing the strains to ciprofloxacin (20.3%). 16.7% of all the strains were resistant to gentamicin. Low levels of resistance were observed by testing the strains to trimethoprim/sulfamethoxazole (3.9%). All the S. aureus strains were sensitive to glycopeptides and the second line anti-Staphylococcal antibiotics: linezolid, quinupristin/dalfopristin.

Conclusion: The majorities of the isolated strains were MSSA and exhibited higher percentage susceptibility, in contrast to those that had been isolated in 2015. Therefore, the policy that was adapted to avoid the spread of resistant strains was effective and recommended the need to introduce the antibioticotherapy control program in the Romanian hospitals.

Biography:

D L Savithramma has completed her PhD from University of Agricultural Sciences, Bangalore, India and Post-doctoral studies as a Biotechnology National Associate from Indian Institute of Science, Bangalore, India. She is a Professor and University Head of Genetics and Plant Breeding at University of Agricultural Sciences, Bangalore, India. She has guided 13 PhD students and 30 Master’s students in Genetics and Plant Breeding. She has published more than 70 papers in reputed journals and has released eight varieties in vegetable cowpea, seed cowpea, peanut and Chrysanthemum.

Abstract:

Tomato is an important commercial vegetable crop. However, tomato is affected by both biotic and abiotic stresses, most importantly; the increasing occurrence of drought worldwide has highlighted the need for development of drought tolerant genotypes in tomato. Identification and selection of genotypes with improved drought tolerance will play an important role in developing tomato genotypes with better yield. The aim of this study was to investigate the effects of drought stress on fruit yield through physiological traits related to drought tolerance viz., SPAD chlorophyll Meter Reading (SCMR), Specific Leaf Area (SLA), Relative water content (RWC) and leaf rolling (LR) and also establish marker association with physiological and fruit yield traits. One hundred (100) germplasm accessions of six cultivated tomato and related species along with three check varieties were evaluated during summer 2014 and 2015 under well watered and water stress condition (stress was imposed on 60^th day after transplanting for a period of 20 days). Population structure was determined using model based method by structure, Neighbor-joining method, Q-matrix population structure and Delta K analysis and all of them separated population into four clusters. Marker-trait associations are established using 145 published SSR markers with syntenic Linkage Disequilibrium (LD) values (r2=0.09) which depicted that some markers detected as most powerful due to high R2 value. Marker SSR 52 associated with SCMR and TGS 2002 for fruit yield per plant and clusters per plant in lycopersicum species. The markers SSR 14 and LELEUZIP for LR, LEet 004 for plant height, fruit yield per plant and LR in cherry species. The marker SSR 593 for days to 50% flowering, SSR 27 for days to first fruit set, SSR 218 for stem girth, TGS 0412 for clusters per plant, SSR 128 for fruits per plant, SSR 599 and SSR 593 for fruit yield in all the spp. This study demonstrated that association mapping in cultivated and related species can enhance the information from QTL studies towards the implementation of marker-assisted selection. From the present study five germplasm accessions namely LA 2976, WIR 13708, EC 676809, EC 677123 and EC 771596 were identified as top drought tolerant genotypes which may be used as parents in hybridization program to develop drought tolerant and high yielding varieties in tomato.