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      Criteria for selecting PEGylation sites on proteins for higher thermodynamic and proteolytic stability.

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          Abstract

          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.

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          Author and article information

          Journal
          J. Am. Chem. Soc.
          Journal of the American Chemical Society
          American Chemical Society (ACS)
          1520-5126
          0002-7863
          Dec 17 2014
          : 136
          : 50
          Affiliations
          [1 ] Department of Chemistry and Biochemistry, Brigham Young University , Provo, Utah 84602, United States.
          Article
          10.1021/ja5095183
          25409346
          75f64605-3bef-4487-bd96-5115f1183e20
          History

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