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Fifteenth Annual
Choosing, Designing and Optimizing Hosts and Platforms 
Methods, Strategies & Applications 
January 24-25, 2013 
 

  

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The demand for larger amounts of protein, in shorter periods of time continues to plague protein researchers. This conference will explore the newest data and innovations relating to the best choices in hosts/systems, as well as ways to “rescue” existing systems and make them work more effectively to produce the quantity and quality of the desired protein.

THURSDAY, JANUARY 24

1:00-2:00 pm Conference Registration

 

Solutions for Difficult Proteins 

1:45 Chairperson’s Opening Remarks

Julio Camarero, Ph.D., Associate Professor, Departments of Pharmacology, Pharmaceutical Sciences and Chemistry, University of Southern California  

1:50 Improving Membrane Protein Expression in E. coli

François Baneyx, Ph.D., Charles W.H. Matthaei Professor, Department of Chemical Engineering, University of Washington

Functional membrane proteins are notoriously difficult to overproduce in native lipid environments because they routinely fail to reach, partition and/or fold into the membrane, causing cellular toxicity. Here, I will describe how the network of molecular chaperones, foldases, translocases and insertases responsible for the biogenesis of inner membrane a-helical and outer membrane b-barrel proteins can be tuned to improve recombinant membrane protein productivity in E. coli.

2:35 New Strategies for the Production of Functional Membrane Proteins Using E. coli as an Expression Host

Jan Kubicek, Ph.D., CSSB-FZJ Institute, Hamburg

Progress in expression of membrane proteins is slowed by the lack of methods and procedures for expression and isolation of these proteins in quantities which allow their functional characterization and structural analysis. We have developed a set of standard protocols for expression and isolation of functional human membrane proteins such as transporters and GPCR’s. We will show their functional characterization and initial data towards structure determination by X-Ray crystallization.

3:05 Using the Skp Chaperone to Increase Recombinant Protein Yields

Jennifer A. Maynard, Ph.D., Departments of Chemistry, Microbiology, and Chemical Engineering, University of Texas at Austin

The periplasmic Skp chaperone has been characterized for its role in outer membrane protein biogenesis, but is increasingly recognized for assisting soluble protein folding. Using a panel of scFvs, we performed in vivo expression, in vitro folding and aggregation assays in the presence or absence of Skp. The results suggest Skp is a holdase, sequestering long-lived intermediates to prevent aggregation.

Life_Technologies3:35 Enhanced Protein Expression using the Expi293TM High Density Expression System

Lester Gutshall, Scientist, Molecular & Protein Biosciences, Janssen R&D

The Expi293™ Expression System is a major advancement in transient expression technology for rapid and ultra high-yield protein production from mammalian cells. As a beta tester, we compared it to our routine transient expression process and we received ~5x higher protein yield resulting in decreased cost per mg protein.

3:50 Refreshment Break in the Exhibit Hall with Poster Viewing

4:30 Production Platforms for Biotherapeutic Glycoproteins: Occurrence, Impact, and Challenges of Non-Human Sialylation

Darius Ghaderi, Ph.D., Senior Research Scientist, Sialix, Inc.

Optimal glycosylation is critical for therapeutic glycoproteins, as glycans can influence their yield, immunogenicity and efficacy. Therapeutic glycoproteins produced in non-human cell lines can differ from human glycans by presenting the terminal Galα1-3Gal (alpha-Gal) modification and N-glycolylneuraminic acid (Neu5Gc). All humans spontaneously express antibodies against both of these glycan structures, risking increased immunogenicity of biotherapeutics carrying such non-human glycan epitopes.

5:00 Efficient Biosynthesis of Native Cyclotides Using Intein-Mediated Protein Trans-Splicing

Julio A. Camarero, Ph.D., Associate Professor, Departments of Pharmacology, Pharmaceutical Sciences and Chemistry, University of Southern California

We report a novel method for the biosynthesis of natively-folded MCoTI-II inside live E. coli cells using a split protein splicing unit. Biosynthesis of genetically encoded cyclotide-based libraries opens the possibility of using single cells as microfactories where the biosynthesis and screening of particular inhibitor can take place in a single process within the same cellular cytoplasm. The cyclotide scaffold has a tremendous potential for the development of therapeutic leads based on their extraordinary stability and potential for grafting applications. We will also report the design and biosynthesis of a MCoTI-grafted cyclotide with the ability to target intracellular and extracellular protein-protein interactions.

5:30 A Robust and Rapid Method of Producing Soluble, Stable and Functional G-Protein Coupled Receptors

Karolina Corin, Researcher, Schauer Laboratory, Center for Biomedical Engineering, Massachusetts Institute of Technology

 

6:00 End of Day



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