Sg. Burton et al., PRODUCTION OF ENANTIOMERICALLY PURE AMINO-ACIDS - CHARACTERIZATION OFSOUTH-AFRICAN HYDANTOINASES AND HYDANTOINASE-PRODUCING BACTERIA, Journal of molecular catalysis. B, Enzymatic, 5(1-4), 1998, pp. 301-305
Chiral amino acid derivatives can be synthesised by the biocatalytic c
onversion of substituted hydantoins using microbial enzymes or resting
cells: a hydantoinase performs the ring-opening cleavage of the hydan
toin to produce an N-carbamylamino acid and N-carbamylamidohydrolase t
hen converts this intermediate to the amino acid, ammonia and CO2. The
hydantoinases from four locally isolated bacterial strains are curren
tly being characterised in terms of conditions for optimal enzyme acti
vity assay, and to demonstrate the effects of pH, temperature, metal i
ons, protease inhibitors, surfactants, and anti-oxidants on hydantoina
se activity in crude extracts. Typically, pH 8, 50 degrees C, and 0.3
mM Cu2+ were found to be optimal. Disruption of cells using a detergen
t or membrane freeze-fracture resulted in increased activities, sugges
ting that the hydantoinase enzymes may be membrane bound. It was also
found that three Pseudomonas strains exhibited higher activities than
the Agrobacterium strain, in terms of hydantoin conversion, with % con
version of hydantoins to N-carbamylamino acids from 66% to 2%. Compari
sons of hydantoinase and amidohydrolase activity in resting cells and
in cell extracts also show marked differences in activity profile for
different strains, e.g., strain RU-KM1 exhibited hydantoinase activity
in whale cells and cell extracts, but amidohydrolase activity only in
cell extracts, while RU-OR showed higher amidohydrolase activity than
hydantoinase activity. (C) 1998 Elsevier Science B.V. All rights rese
rved.