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        Molecular Bioprocessing Research
 
 

 

 

 

  

The Journal of Physical Chemistry B

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"Molecular Simulations of Multimodal Ligand-Protein Binding: Elucidation of Binding Sites and Correlation with Experiments"

Freed, AS, Garde S, Cramer SM

2011, 115 (45), pp 13320-13327

B&B cover image

Biotechnology and Bioengineering

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"Investigation of Protein Binding Affinity and Preferred Orientations in Ion Exchange Systems Using a Homologous Protein Library"

Chung WK, Hou Y, Freed A, Holstein M, Makhatadze GI, Cramer SM

2009, 102 (3), pp 869-881

 
 

        About Dr. Cramer

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Professor Steven Cramer is the William Weightman Walker Professor of Polymer Engineering at Rensselaer Polytechnic Institute, Troy, New York. He is currently conducting research on several areas related to protein-surface interactions and molecular bioprocessing. In addition, Professor Cramer is known worldwide for his expertise in separations in general. He served as the Editor-in-Chief of the International journal Separations, Science and Technology for 20 years. Professor Cramer was the awarded the Alan S. Michaels Award for the Recovery of Biological Products (ACS Division of Biochemical Technology) and the 2016 ACS National Award in Separation Science and Technology. He was also awarded Rensselaer’s School of Engineering Outstanding Professor Award and the Research Excellence Award. Dr. Cramer was given a Presidential Young Investigator award from the National Science Foundation, the Early Career Award from Rensselaer Polytechnic Institute as well as several teaching awards. Professor Cramer has been elected a fellow of the American Association for the Advancement of Science, American Institute of Chemical Engineers, the American Chemical Society and the American Institute for Medical and Biological Engineering. He has chaired several prestigious meetings including 2 International HIC/RPC Bioseparation Conferences, 2 ACS Recovery of Biological Products Meetings and the Gordon Conference on Reactive Polymers. Prof. Cramer is a consultant for several biopharmaceutical and bioseparation companies. He is also serving on the FDA panel for biosimilars and is the Chair of the Recovery of Biological Products Board. Prof. Cramer has published over 185 papers in peer-reviewed journals and has 11 patents. Importantly, he has produced 45 Ph.D. students who have gone on to leadership positions in the biotechnology industry and academia.

      Our Research

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The research in the Cramer laboratory involves state of the art experimental and theoretical investigations into novel bioseparations systems. Topics include: prediction of protein binding affinity and multiscale modeling of chromatographic systems, design of chemically selective displacers, development of efficient antibody separation systems, fundamental studies in multimodal chromatography, novel lab on chip separations systems, protein unfolding in chromatographic systems, chemometrics for process analytical technology, multilevel automated peptide synthesis/screening system for design of affinity peptides and biosensors, smart biopolymer affinity precipitation systems, use of high throughput screening for quality by design of large scale bioprocesses, design of novel affinity reagents for purification of therapeutic enzymes, and development of hierarchical nano-bio materials for bioprocessing. As one example, recent work that combines protein libraries, NMR studies, molecular dynamics simulations and high throughput chromatographic investigations has provided significant insight into the design of a new generation of mixed mode chromatographic systems.

Recent Highlights:

Journal of Chromatography A
Domain contributions to antibody retention in multimodal chromatography systems 
Robinson, J., Roush, D., Cramer, S.M.
2018, 1563, 89-98
Abstract: Although a platform process has been established for purification of antibodies, a deep, fundamental understanding of how these molecules interact with chromatography resins has yet to be developed. The increasing prevalence of antibody-related therapeutics and associated purification challenges further motivate research into these molecular level interactions. The objective of this work is to understand the nature (i.e. size and properties) of preferred protein-ligand binding regions for large, multi-domain molecules such as antibodies. In this work, three antibodies with pI 7.5–8.3 and varying hydrophobicity were enzymatically digested to create (Fab)2, Fab, and FC fragments. Linear salt gradient chromatography experiments from 0 to 1M NaCl were carried out with the full mAbs and the fragments in several multimodal chromatography systems at pH 6. The retention of the constituent fragments was then compared to that of the mAb to gain insight into the relative importance of these different domains and the contribution of each domain to the binding of the full mAb in these systems. While some mAbs were dominated by contribution from the FC constant region, others were primarily driven by the (Fab)2 interactions. The domain contributions for each mAb were connected to the unique distribution of surface charge and hydrophobicity using protein surface property maps. This work lays the foundation for identifying the key surface patches on large, multi-domain molecules that are important interaction sites in various multimodal systems. Finally, this work has important implications for the separation of product related variants as well as the design of complex therapeutics for biomanufacturability.

Biotechnology and Bioengineering
An impurity characterization based approach for the rapid development of integrated downstream purification processes 
Timmick, S.M., Vecchiarello, N., Goodwine, C., Crowell, L.E., Love, K.R., Love, J.C., and Cramer, S.M.
2018, 119,2048-2060
Abstract: In this study, we describe a new approach for the characterization of process‐related impurities along with an in silico tool to generate orthogonal, integrated downstream purification processes for biological products. A one‐time characterization of process‐related impurities from product expression in Pichia pastoris was first carried out using linear salt and pH gradients on a library of multimodal, salt‐tolerant, and hydrophobic charge induction chromatographic resins. The Reversed‐phase ultra‐performance liquid chromatography (UPLC) analysis of the fractions from these gradients was then used to generate large data sets of impurity profiles. A retention database of the biological product was also generated using the same linear salt and pH gradients on these resins, without fraction collection. The resulting two data sets were then analyzed using an in silico tool, which incorporated integrated manufacturing constraints to generate and rank potential three‐step purification sequences based on their predicted purification performance as well as whole‐process “orthogonality” for impurity removal. Highly ranked sequences were further examined to identify templates for process development. The efficacy of this approach was successfully demonstrated for the rapid development of robust integrated processes for human growth hormone and granulocyte‐colony stimulating factor.

 

 

   

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