title: Protein-polymer conjugation via ligand affinity and photoactivation of glutathione S-transferase.
abstract: A photoactivated, site-selective conjugation of poly(ethylene glycol) (PEG) to the glutathione (GSH) binding pocket of glutathione S-transferase (GST) is described. To achieve this, a GSH analogue (GSH-BP) was designed and chemically synthesized with three functionalities: (1) the binding affinity of GSH to GST, (2) a free thiol for polymer functionalization, and (3) a photoreactive benzophenone (BP) component. Different molecular weights (2 kDa, 5 kDa, and 20 kDa) of GSH-BP modified PEGs (GSBP-PEGs) were synthesized and showed conjugation efficiencies between 52% and 76% to GST. Diazirine (DA) PEG were also prepared but gave conjugation yields lower than for GSBP-PEGs. PEGs with different end-groups were also synthesized to validate the importance of each component in the end-group design. End-groups included glutathione (GS-PEG) and benzophenone (BP-PEG). Results showed that both GSH and BP were crucial for successful conjugation to GST. In addition, conjugations of 5 kDa GSBP-PEG to different proteins were investigated, including bovine serum albumin (BSA), lysozyme (Lyz), ubiquitin (Ubq), and GST-fused ubiquitin (GST-Ubq) to ensure specific binding to GST. By combining noncovalent and covalent interactions, we have developed a new phototriggered protein-polymer conjugation method that is generally applicable to GST-fusion proteins.
authors.affiliation: Department of Chemistry & Biochemistry and the California NanoSystems Institute, University of California, Los Angeles , 607 Charles E. Young Drive East, Los Angeles, California 90095, United States.
authors.forename: En-Wei
authors.initials: EW
authors.lastname: Lin
authors.forename: Natalie
authors.initials: N
authors.lastname: Boehnke
authors.forename: Heather D
authors.initials: HD
authors.lastname: Maynard
mesh_terms: D000818:Animals; D001577:Benzophenones / Q000737:chemistry*; D005978:Glutathione / Q000031:analogs & derivatives*; D005982:Glutathione Transferase / Q000737:chemistry*; D006736:Horses; D008024:Ligands; D008027:Light; D011092:Polyethylene Glycols / Q000737:chemistry*
chemical_list: D001577:Benzophenones; D008024:Ligands; D011092:Polyethylene Glycols; C047723:benzophenone; D005982:Glutathione Transferase; D005978:Glutathione