Criteria for Selecting PEGylation Sites on Proteins for Higher Thermodynamic and Proteolytic Stability
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- 作者：Paul B. Lawrence ; Yulian Gavrilov ; Sam S. Matthews ; Minnie I. Langlois ; Dalit Shental-Bechor ; Harry M. Greenblatt ; Brijesh K. Pandey ; Mason S. Smith ; Ryan Paxman ; Chad D. Torgerson ; Jacob P. Merrell ; Cameron C. Ritz ; Maxim B. Prigozhin ; Yaakov Levy ; Joshua L. Price
- 刊名：Journal of the American Chemical Society
- 出版年：2014
- 出版时间：December 17, 2014
- 年：2014
- 卷：136
- 期：50
- 页码：17547-17560
- 全文大小：756K
- ISSN：1520-5126
文摘
PEGylation of protein side chains has been used for more than 30 years to enhance the pharmacokinetic properties of protein drugs. However, there are no structure- or sequence-based guidelines for selecting sites that provide optimal PEG-based pharmacokinetic enhancement with minimal losses to biological activity. We hypothesize that globally optimal PEGylation sites are characterized by the ability of the PEG oligomer to increase protein conformational stability; however, the current understanding of how PEG influences the conformational stability of proteins is incomplete. Here we use the WW domain of the human protein Pin 1 (WW) as a model system to probe the impact of PEG on protein conformational stability. Using a combination of experimental and theoretical approaches, we develop a structure-based method for predicting which sites within WW are most likely to experience PEG-based stabilization, and we show that this method correctly predicts the location of a stabilizing PEGylation site within the chicken Src SH3 domain. PEG-based stabilization in WW is associated with enhanced resistance to proteolysis, is entropic in origin, and likely involves disruption by PEG of the network of hydrogen-bound solvent molecules that surround the protein. Our results highlight the possibility of using modern site-specific PEGylation techniques to install PEG oligomers at predetermined locations where PEG will provide optimal increases in conformational and proteolytic stability.