Binding of by-product analog benzylsuccinic acid by carboxypeptidase A

A variety of carboxylic and dicarboxylic acids were examined as inhibitors of bovine carboxypeptidase A. Of these, the most effective was 2(R)-benzyl-3-carboxypropionic acid (the L isomer of benzylsuccinic acid). This inhibitor appeared to be purely competitive with respect to hydrolysis of both esters and peptides, and exhibited Ki = 4.5 (± 0.8) × 10-7 M when assayed against a variety of substrates at pH 7.5 in 0.5 M NaCl at 25°. The stoichiometry, site, and pH dependence of binding were examined; the results suggest that a single molecule of a monoanionic species of L-benzylsuccinic acid is bound, and that this occurs at the active site. Binding is associated with detectable changes in ultraviolet spectra, resistance of the enzyme to various kinds of irreversible inactivation, an increase in enzyme solubility, and a change in the sites of enzyme derivatization with tetranitromethane. Kinetic analysis of enzyme inhibition by the products hippuric acid and L-phenylalanine suggests that binding of one of these products does not appreciably affect binding of the other. It is suggested that L-benzylsuccinate resembles the collected products of peptide hydrolysis, and is thus bound with an affinity resembling their combined affinity. Binding of by-product analog benzylsuccinic acid by carboxypeptidase A.

Economical succinic acid production from cane molasses by Actinobacillus succinogenes

In this work, production of succinic acid by Actinobacillus succinogenes CGMCC1593 using cane molasses as a low cost carbon source was developed. In anaerobic bottles fermentation, succinic acid concentration of 50.6+/-0.9 g l(-1) was attained at 60 h using an optimum medium containing molasses pretreated with sulfuric acid, resulting in a yield of 79.5+/-1.1% and sugar utilization of 97.1+/-0.6%. When batch fermentation was carried out in a 5-l stirred bioreactor with pretreated molasses, 46.4 g l(-1) of succinic acid was attained at 48 h and faster cells growth was also observed. Fed batch fermentation was performed to minimize the substrate (sugar) inhibition effect, giving 55.2 g l(-1) of succinic acid and 1.15 g l(-1)h(-1) of productivity at 48 h. The present study suggests that the inexpensive cane molasses could be utilized for the economical and efficient production of succinic acid by A. succinogenes.