Investigations on enzyme-catalyzed peptide synthesis

Abstract

Enzymes have been found to be catalytically active in organic solvents. Chymotrypsin was used to synthesize a wide range of peptides when suspended in organic solvents. This method overcame such problems as secondary hydrolysis and poor solubility in aqueous mixtures, and allowed processes to occur that were impossible in water. Syntheses involving D-amino acid derivatives were possible under these conditions. Molecular modelling studies have been carried out and structure-reactivity relationships have been drawn by using hydrolytic reactions catalyzed by chymotrypsin suspended in organic solvents with low water content. Kinetic studies of chymotrypsin suspended in organic solvents have shown that the enzyme does not have a classical Michaelis- Menten behaviour, but shows cooperative effects with respect to the binding of the substrates. The role of the essential water has been investigated. The use of the different chymotrysins in peptide synthesis has been investigated and 7 -chymotrypsin, inactive in water, has been found fully active when suspended in organic solvents. The enzymatic synthesis of leucine-enkephalinamide was carried out and tandem mass spectrometry was used to determine the composition of the reaction mixture. The last coupling step used an enzyme in an organic solvent. Potential competitive inhibitors of proteases have been designed and synthesized. Several methods for the synthesis of 3-keto esters were investigated and diastereoselective reductions of 3- keto esters derived from amino acids were achieved