Authors: Maullu, Carlo; Raimondo, Domenico; Caboi, Francesca¹; Giorgetti, Alejandro; Sergi, Mauro¹; Valentini, Maria²; Tonon, Giancarlo¹; Tramontano, Anna

Source: FEBS Journal, Volume 276, Number 22, November 2009 , pp. 6741-6750(10)

Publisher: Wiley-Blackwell

Abstract:

Poly(ethylene glycol) (PEG) is a widely used polymer employed to increase the circulating half-life of proteins in blood and to decrease their immunogenicity and antigenicity. PEG attaches to free amines, typically at lysine residues or at the N-terminal amino acid. This lack of selectivity can present problems when a PEGylated protein therapeutic is being developed, because predictability of activity and manufacturing reproducibility are needed for regulatory approval. Enzymatic modification of proteins is one route to overcome this limitation. Bacterial transglutaminases are enzyme candidates for site-specific modification, but they also have rather broad specificity. The need arises to be able to predict a priori potential PEGylation sites on the protein of interest and, especially, to be able to design mutants where unique PEGylation sites can be introduced when needed. We investigated the feasibility of a computational approach to the problem, using human granulocyte colony-stimulating factor as a test case. The selected protein is therapeutically relevant and represents a challenging problem, as it contains 17 potential PEGylation sites. Our results show that a combination of computational methods allows the identification of the specific glutamines that are substrates for enzymatic PEGylation by a microbial transglutaminase, and that it is possible to rationally modify the protein and introduce PEG moieties at desired sites, thus allowing the selection of regions that are unlikely to interfere with the biological activity of a therapeutic protein.

Keywords: molecular dynamics; PEGylation; protein-protein docking; site-directed mutagenesis; transglutamination

Document Type: Research article

DOI: http://dx.doi.org/10.1111/j.1742-4658.2009.07387.x

Affiliations: 1:  Bio-Ker S.r.l., c/o Sardegna Ricerche Scientific Park, Pula, Cagliari, Italy 2:  CRS4-Bioinformatics Laboratory, c/o Sardegna Ricerche Scientific Park, Pula, Cagliari, Italy

Publication date: 2009-11-01

Related content

• In this: publication
• By this: publisher
• In this Subject: Anatomy & Physiology ,  Biology ,  Chemistry (General) ,  Biochemistry
• By this author: Maullu, Carlo ;  Raimondo, Domenico ;  Caboi, Francesca ;  Giorgetti, Alejandro ;  Sergi, Mauro ;  Valentini, Maria ;  Tonon, Giancarlo ;  Tramontano, Anna

Website © Publishing Technology. Article copyright remains with the publisher, society or author(s) as specified within the article.

• Terms and conditions
• Privacy policy
• Information for advertisers