SYNTHESIS OF POLY(ETHYLENE GLYCOL) DERIVATIVES WITH DIFFERENT BRANCHINGS AND THEIR USE FOR PROTEIN MODIFICATION

Monomethoxy linear poly(ethylene glycol) (PEG) with a terminal hydroxy group was coupled to monobromoacetic acid, protocatechnic acid and gallic acid to synthesize one branched (PEG1), two branched (PEG2) and three branched PEG derivatives (PEG3) each having only one carboxyl group in a molecule. The PEG derivatives were chemically fixed to trypsin through amidation with its amino groups using the PEG carboxyl group. The PEG-modified trypsins with different degrees of modification were subjected to three enzymatic reactions. When casein hydrolysis and trypsin autolysis were performed using the PEG-modified trypsins, both of the enzymatic reactions were strongly suppressed with the PEG modification. On the other hand, inhibition of trypsin activity by trypsin inhibitor was scarcely affected by the PEG modification, whereas trypsin digestion by pepsin was greatly protected by the PEG modification in the order of PEG3 > PEG2 > PEG1. All these results could be consistently explained in terms of steric hindrance brought about by fixation of the PEG chains on the trypsin molecule.