Haruyuki Ohkishi

Synthesis of D-cysteine from 3-chloro-D-alanine and hydrogen sulfide by 3-chloro-D-alanine hydrogen chloride-lyase (deaminating) of Pseudomonasputida

Abstract Synthesis of D-cysteine from 3-chloro-D-alanine and hydrogen sulfide is catalyzed by highly purified 3-chloro-D-alanine hydrogen chloride-lyase from Pseudomonas putida . The synthetic reaction proceeds optimally at pH 8.5, as a function of enzyme concentration and incubation time. The enzymatically synthesized D-cysteine was isolated from the large scale reaction mixture and identified by physicochemical means.

Synthesis of D-cysteine-related amino acids by 3-chloro-D-alanine chloride-lyase of Pseudomonasputida CR 1-1

Using the beta-replacement reaction of 3-chloro-D-alanine chloride-lyase from Pseudomonas putida CR 1-1, S-methyl-, S-ethyl-, S-n-propyl-, S-n-butyl-, S-phenyl-, S-benzyl-, S-(2,3-dihydroxypropyl)- and S-(2-hydroxyethyl)-D-cysteine were synthesized from 3-chloro-D-alanine and methyl-, ethyl-, n-propyl-, n-butyl-, phenyl-, benzyl-mercaptans, alpha-thioglycerol and 2-mercaptoethanol, respectively. The enzymatically synthesized D-cysteine-related amino acids were isolated from the large scale reaction mixture and identified physicochemically.

Synthesis of d-Cysteine from a Racemate of 3-Chloroalanine by Phenylhydrazine-treated Cells of Pseudomonas putida CR 1-1

3-Chloro-d-alanine chloride-lyase, which occurs in the cells of Pseudomonas putida CR 1-1, catalyzes the β-replacement reaction of 3-chloro-d-alanine in the presence of a high concentration of sodium hydrosulfide to form d-cysteine. Using the β-replacement reaction, the synthesis of d- cysteine from a racemate of 3-chloroalanine by resting cells was investigated. In order to depress the synthesis of l-cysteine from 3-chloro-l-alanine and the enzymatic degradation of 3-chloro-l- alanine, treatment of the cells with 5mM phenylhydrazine was utilized. Under the optimal conditions, 94% of the added 3-chloro-d-alanine could be converted to d-cysteine and 81% of the added 3-chloro-l-alanine was recovered.