Role of Lysine in catalysing Ester Hydrolysis
RECENTLY, Gero and Withrow1 have cited the catalysis of alkaline triglyceride hydrolysis by lysine as a chemical model of a biological reaction. They postulated a bifunctional2 attack by lysine upon the carbonyl O and C of the ester linkage as the mechanism of this hydrolysis.
Kinetics of aldose-amino acid interaction.
Abstract 1. 1. The kinetics of the reaction between various aldoses and amino acids and peptides has been investigated. 2. 2. A method for the determination of rate constants, based on the addition of alkali to keep the pH of the reaction mixture constant, has been developed. 3. 3. The velocity of the reaction decreases with increase of pH according to the equation dRG − d t = R − .G kα + kβ .OH − − RG − (kα + kβ .OH − )L where RG − is the concentration of the reaction product, R − —that of the amino acid anion, G—of the aldose; OH − —the hydroxyl ion concentration and k α , k β pH independent rate constants. 4. 4. The rate constant k β is inversely proportional to the rate constant of the primary attack of the amino acid on the aldose. The rate of the primary reaction increases with the basicity of the amino component in accordance with the Hammett relation.
New method for analysis of enzymatic reactions: Thermolysin-catalyzed synthesis of aspartame precursor
Aspartame is an artificial sweetener which is 200 times sweeter than sucrose. A precursor of aspartame, N-(benzyloxycarbonyl)-l-aspartyl-l-phenylalanine methyl ester (Z-APM) was synthesized and hydrolyzed with thermolysin in an aqueous monophasic system. Graphical plotting methods and/or nonlinear regression have been used to analyze enzymatic reactions because of the nonlinearity of enzymatic rate equations. There are, however, problems: the former require accurate measurement of the initial reaction rate and the latter gives no analytical information. The authors proposed a method to solve the nonlinear enzymatic rate equations with respect to the concentration of a product with the Taylor expansion, especially for the reversible Theorell-Chance mechanism (Murakami, Y., Hirata, M., and Hirata, A., J. Ferment. Bioeng., 82, 246–252, 1996). This solution, however, is complex for practical use. In this paper, a simpler solution to explain the thermolysin-catalyzed synthesis of Z-APM is obtained. New methods to determine the kinetic parameters, especially for the reversible Theorell-Chance mechanism, based on the approximated solution are proposed. The results calculated using the obtained kinetic parameters can satisfactorily explain the experimental results, which demonstrates the effectiveness of the proposed analysis. We must know the relation between the concentrations of substrates and a product, and accurately evaluate of the result obtained using Taylor expansion when we determine the kinetic parameters using our newly developed methods. The methods to determine the kinetic parameters of the reversible Theorell-Chance mechanism described in this paper is, however, expected to be applied to general enzymatic reactions.