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Overview | Short
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WEDNESDAY, JANUARY 14
7:00am – 5:30pm Registration
7:30 Breakfast Workshop (Sponsorship Available)
PROTEIN EXPRESSION WITH A DOWNSTREAM FOCUS
8:15 Chairperson’s Remarks
Dennis M. Kraichely, Ph.D., Centocor R & D, Inc.
8:20 Codon Harmonization Leads to Enhanced Heterologous Protein Expression
Evelina Angov, Ph.D., Chief, Laboratory Molecular Parasitology USMMVP-WRAIR
For Plasmodium falciparum, codon harmonization has yielded dramatic improvements in plasmid stability, overall expression levels and soluble protein yield (indicator of correct folding) compared to expression by the non-recoded native genes in E. coli. Since adjustments to relative codon usage improve the reliability of functional protein expression, this approach may represent a paradigm shift for heterologous protein expression, with important consequences for both structural biology and biotechnology.
8:50 Transgenic Plant Systems for the Production of Structural Proteins
Julio A. Báez, Ph.D., Senior Research Fellow, FibroGen USA
Plant-based transgenic systems could offer a cost effective system to deliver thousands of metric tons per year by producing recombinant structural proteins as by-products of biorefining. Data describing the expression of recombinant collagen-related proteins in transgenic corn and barley seeds will be discussed indicating the feasibility of using these plant-based transgenic production systems for making animal-component free structural proteins with consistent composition and structure.
9:20 Automated Small-Scale Transfections to Scale Up Lead Molecule
Matthew Husovsky, Senior Associate Scientist, Protein Sciences, Centocor, A Johnson & Johnson Company
We have a rapid system that can go from screening clones to characterizing lead molecules in less than 4 months. By automating 96-well DNA production, along with multi-well transfections, we have the potential to screen thousands of variants per week. We are working with a robust transient transfection system that allows us to express and purify up to 40 lead candidates in the 5mg scale in one-week. We are using large-scale transient CHO transfections to characterize 2-8 lead molecules, and generating 50mg of each protein prior to NME declaration.
9:50 Coffee Break in the Exhibit Hall
10:45 Post-Transformational Vector Amplification in the Yeast Pichia pastoris
James M. Cregg, Ph.D., Keck Graduate Institute of Applied Life Sciences
11:15 Development of a Protease Production Platform for Structure-Based Drug Design
Edward Fox, Ph.D., Vertex Pharmaceuticals
11:45 Panel Discussion: Looking Downstream Early to Prevent Problems Later
| 12:15
Pfēnex Expression
TechnologyTM , The New Paradigm of Microbial Strain Development for Biopharmaceutical Production |
Sponsored by
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Charles
S. Squires, Ph.D., Senior Director of R&D, Dowpharma
The P. fluorescens expression platform, Pfēnex Expression TechnologyTM, has been engineered to deliver host strains expressing large amounts of high quality target protein within very short development times. This has been done by creating an off the shelf toolbox of expression plasmids and host strains. Together with, rapid, small scale (96 well) growth and assay techniques, hundreds of unique expression strains are tested in parallel to find combinations of expression strategy and host cell phenotype producing the highest amount of best quality product. Further, methods have been developed to very rapidly test multiple fermentation conditions on selected strains in scaled-down, but scalable bioreactors. Application of these new tools allows identification of a production strain and development of a first fermentation process in less than 10 weeks from initiation of gene cloning. This presentation will describe the development and operation of the platform as well as case studies highlighting its advantages |
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| 12:30 Luncheon Workshop
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Sponsored by
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Streamlining
Fragment-Based Drug Discovery Using a Panel of Protein Kinase Crystals
Faming Zhang,
Ph.D., President, Crown Biosciences
Using a kinase and phosphotase co-expression system, a diverse
protein kinase crystal panel has been generated. A streamlined
process to determine binding affinity, co-crystallization and
structure refinement process has been created to increase the
throughput of fragment-based lead generation for protein kinases.
Some case studies of the kinase specificity generated from different
fragment inhibitor precursors will be discussed. |
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CASE STUDIES
1:45 Chairperson’s Remarks
Dennis M. Kraichely, Ph.D., Centocor R & D, Inc.
1:50 Expression and Purification of the Functional
Domain of DKK2 in E. coli
Jie Zheng, Ph.D., Associate Member, Structural Biology, St. Jude Children's Research Hospital
Dickkopf (Dkk) proteins inhibit the canonical Wnt signaling pathway. In this study, we developed a protocol to express the C-terminal domain in E. coli and to purify it through chromatography. We demonstrated that this protocol allows us to effectively generate s highly purified and fully activated protein for structural biology studies. Therefore, this protocol represents an important advancement that will enable more efficient research into the structural and functional properties of DKK proteins.
2:20 In vivo Biotinylation of Recombinant Proteins in the Protozoan Host Leishmania tarentolae
Zoltán Konthur, Ph.D., Group Leader - In Vitro Ligand Screening, Department of Vertebrate Genomics, Max Planck Institute for Molecular Genetics
2:50 Technology Watch
REEMAC: An Platform for Recombinant Protein Production in Cultured Mammalian Cells
Qiang Li, Ph.D., Chief Scientific Officer, ATGCell, ATGCell Inc.
We developed an exogenous expression platform REEMAC to quickly and efficiently produce recombinant proteins from cultured mammalian cells by creating a set of new expression vectors and utilization of our proprietary ExpressBoost reagents. In this platform, we examined a variety of expression elements and optimized the entire expression cassette structure. We also formulated a set of chemical cocktails for further expression enhancement. We have applied REEMAC to express over 100 target proteins, including cytokines, growth factors, hormones, cell surface antigen receptors, proteases and monoclonal antibodies in CHO and HEK cells.
3:05 Consistent Development of Stable-High Expressing Mammalian Cell Lines in Four Months Using GPEx®
Greg Bleck, Ph.D., Senior Director Cell Line Engineering, Gala Biotech, A Catalent Pharma Solutions Company
The GPEx® method of cell line engineering generates high-expressing, genetically stable cells for all mammalian cell types. Antibiotic selection is not needed as part of the procedure, so multiple gene constructs can be added individually, at different gene ratios without any requirement for antibiotic resistance markers. Specific productivities of GPEx® antibody producing cell lines range from 30 -70 picograms/cell/day without upstream process development, while cell lines producing non-antibody proteins as expected have a much broader range of productivities, but if the protein has no secretion problems and does not inhibit cell growth, specific productivities similar to antibodies are obtained.
| 3:20 Technology Spotlight
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Sponsored by
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Novel Surface Chemistries and Systems for the Label-Free Determination of the Kinetics of Biomolecular Interactions
John Quinn, Ph.D., Chief Science Officer, ICx Technologies
ICx Technologies has developed a range of surface chemistries and systems to assist researchers in determining kinetic and affinity rate constants for a wide range of interactions. Applications utilizing these novel surfaces will be discussed along with some of the unique capabilities of the SensiQ systems. |
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3:35 Refreshment Break in the Exhibit Hall
4:15 Poster Awards in the Exhibit Hall
EMERGING TECHNOLOGY
4:30 An Efficient Screening Method of Libraries for Expression of Soluble and Crystallizable Proteins
and Domains in E. coli
Pär Nordlund, Ph.D., Evitaproteoma AB
Using a library of different protein constructs, the SPOTLIGHT system is able to detect constructs with soluble expression at the colony level. This is performed by applying a separation step through a filter at lysis, followed by immunological detection. The method constitutes an efficient HTP tool for screening for soluble protein expression which requires no automation. The method also reflects the situation at scale-up since detection is performed on lysate and no fusion reporter is used. In addition, it potentially increases the probability of obtaining well-diffracting crystals through its capacity to identify many different constructs in one single experiment.
5:00 Production of (Therapeutic) Non-Enzyme Proteins Using a Versatile PAOX1-Promoter Library of Pichia pastoris
Thomas Purkarthofer, PhD, Head R&D, Biotechnology, VTU Technology
Employing our proprietary promoter library of the strongest promoter known to date, the Alcohol Oxidase 1 promoter (AOX1) from Pichia pastoris, controllable expression for several proteins at once is feasible. Co-expression of e.g. chaperones for correct protein folding along with particular target proteins eventually requires different expression characteristics for the helper and target protein, respectively, which can be achieved employing our PAOX1-library. Apart from several examples of high-level expression of industrial enzymes, we will focus on case studies of successful production of highly relevant (therapeutic) non-enzyme proteins.
5:30 Close of Conference
Overview | Short
Courses | Day 1 |
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For more information, please contact:
Elizabeth Lamb, Conference Producer
Phone: 207-493-7874
E-mail: elamb@healthtech.com
For exhibit and sponsorship information, please contact:
Suzanne Carroll, Manager-Business
Development
Phone: 781-972-5452
E-mail: scarroll@healthtech.com
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