Effects of chemical modification of lysine residues in trypsin

Chemical modifications are a simple method to identify and modify functiona l determinants of enzymes. In the case of serine proteases, it is possible to induce characteristics which are advantageous for peptide synthesis. In this work, we investigated the influence of guanylation and succinylation o f lysine residues on the S'-subsite specificity, the catalytic behavior and stability of trypsin. We have found, that succinylation leads to an about 10-fold better acceptance of basic residues in P1', whereas guanylation sho ws no remarkable effects. Furthermore, guanylation enhances, succinylation reduces the general enzyme-substrate interactions in P2'. The structural fu ndamentals of these specificity changes are discussed. The catalytic behavi or of trypsin was not influenced by guanylation and succinylation but an en hancement of the stability against autolytic processes by introducing addit ional negative charges into the protein was observed. (C) 2000 Elsevier Sci ence B.V. All rights reserved.