Transesterification reactions catalyzed by chymotrypsin.

As recently reported, the behavior of acetyl chymotrypsin with ethanol consists in a rapid transfer of the single acetyl group to that alcohol with the formation of ethyl acetate (1). Thus, the splitting of nitrophenyl acetates by chymotrypsin, faster in ethanolic than in aqueous solutions, may be explained by the formation of acetyl chymotrypsin as an intermediate and its subsequent preferential breakdown with ethanol rather than with water. The net result is of course an enzymatic transfer of acetyl from nitrophenyl acetate to ethanol, and the comparatively rapid breakdown of the acetate may be thought of as contingent upon the presence of the alcohol as a L‘cosubstrate” (2, 3). It was of interest to see whether other substances acted in a manner analogous to ethanol, and a qualitative trial of substances in the laboratory, without any attempt at a thorough survey, showed that most of them did not markedly accelerate the decomposition of p-nitrophenyl acetate by chymotrypsin. It was observed that several substances besides cysteine and glutathione (4) decomposed p-nitrophenyl acetate at pH 6.2 without chymotrypsin, among them ethanolamine, histidine, and glycine, the last in fairly high concentration only. The substances which did accelerate the chymotryptic breakdown of p-nitrophenyl acetate without being active by themselves in this respect happened to be alcohols (in the cases tried), and included glycerol and monoacetin. It therefore seemed worth while to study the behavior of a series of the aliphatic monohydric alcohols as far as their solubility would permit. For practical reasons the series was limited to methanol, ethanol, the two propanols, the four butanols, n-pentanol, and n-hexanol. It was found that the straight chain alcohols accelerated the breakdown of p-nitrophenyl acetate much more rapidly than their branched chain isomers. Moreover, the reaction rates observed with the straight chain compounds increased considerably with the length of the carbon chain (methanol being an exception, as it reacted faster than ethanol). Thus the reaction with n-hexanol was found to be at least fifteen times faster than