Reaction mechanism and stereochemistry of gamma-hexachlorocyclohexane dehydrochlorinase LinA.
gamma-Hexachlorocyclohexane dehydrochlorinase (LinA) catalyzes the initial steps in the biotransformation of the important insecticide gamma-hexachlorocyclohexane (gamma-HCH) by the soil bacterium Sphingomonas paucimobilis UT26. Stereochemical analysis of the reaction products formed during conversion of gamma-HCH by LinA was investigated by GC-MS, NMR, CD, and molecular modeling. The NMR spectra of 1,3,4,5,6-pentachlorocyclohexene (PCCH) produced from gamma-HCH using either enzymatic dehydrochlorination or alkaline dehydrochlorination were compared and found to be identical. Both enantiomers present in the racemate of synthetic gamma-PCCH were converted by LinA, each at a different rate. 1,2,4-trichlorobenzene (1,2,4-TCB) was detected as the only product of the biotransformation of biosynthetic gamma-PCCH. 1,2,4-TCB and 1,2,3-TCB were identified as the dehydrochlorination products of racemic gamma-PCCH. delta-PCCH was detected as the only product of dehydrochlorination of delta-HCH. LinA requires the presence of a 1,2-biaxial HCl pair on a substrate molecule. LinA enantiotopologically differentiates two 1,2-biaxial HCl pairs present on gamma-HCH and gives rise to a single PCCH enantiomer 1,3(R),4(S),5(S),6(R)-PCCH. Furthermore, LinA enantiomerically differentiates 1,3(S),4(R),5(R),6(S)-PCCH and 1,3(R),4(S),5(S),6(R)-PCCH. The proposed mechanism of enzymatic biotransformation of gamma-HCH to 1,2,4-TCB by LinA consists of two 1,2-anti conformationally dependent dehydrochlorinations followed by 1,4-anti dehydrochlorination.
Trantírek
L
L
Laboratory of Biomolecular Structure and Dynamics, Masaryk University, Kotlarska 2, 611 37 Brno, Czech Republic.
Hynková
K
K
Nagata
Y
Y
Murzin
A
A
Ansorgová
A
A
Sklenár
V
V
Damborský
J
J
D001426:Bacterial Proteins / Q000737:chemistry* / Q000502:physiology; D008401:Gas Chromatography-Mass Spectrometry; D001556:Hexachlorocyclohexane / Q000031:analogs & derivatives* / Q000378:metabolism; D008190:Lyases; D009682:Magnetic Resonance Spectroscopy; D008968:Molecular Conformation
D001426:Bacterial Proteins; C010275:pentachlorocyclohexene; D001556:Hexachlorocyclohexane; D008190:Lyases; C071445:linA protein, Pseudomonas paucimobilis