Purification and characterization of an extremely halophilic acetoacetyl-CoA thiolase from a newly isolated Halobacterium strain ZP-6.
The extremely halophilic archaeon ZP-6 was isolated from Ai-Ding salt lake in Xinjiang Uighur Autonomous Region of the People's Republic of China. Based on its physiological properties, 16S rDNA sequence, and DNA-DNA homology with known haloarchaea, the isolate was tentatively identified as a Halobacterium sp. An acetoacetyl-CoA thiolase was purified and characterized from this organism. The native enzyme has a molecular mass of 80 +/- 8 kDa and consists of two identical subunits of 43 +/- 2 kDa each. The N-terminus 14 amino acid residues were sequenced and showed identity with the respective part of a putative thiolase (AcaB1) of Halobacterium sp. NRC-1. The purified enzyme has an optimal pH of 7.9 for acetoacetyl-CoA thiolysis. The thiolytic activity was inhibited by the presence of Mg'- and was stimulated by KCl or NaCl. The thiolysis reaction of Halobacterium sp. ZP-6 thiolase can be inhibited by either substrate when present in excess. The distinct kinetic profile indicates that the thiolase from Halobacterium sp. ZP-6 may have a different catalytic mechanism from the so-called ping-pong mechanism employed by other thiolases. To our knowledge, this is the first report of the purification and characterization of a halophilic thiolase from an archaeal species.
Liu
Tiehan
T
Institute of Microbiology, Chinese Academy of Sciences, Beijing, ROC.
Liu
Shuang-Jiang
SJ
Xue
Yanfen
Y
Ma
Yanhe
Y
Zhou
Peijin
P
D000101:Acetyl-CoA C-Acetyltransferase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D000595:Amino Acid Sequence; D002681:China; D006217:Halobacterium / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D021122:Protein Subunits; D017386:Sequence Homology, Amino Acid; D012965:Sodium Chloride / Q000494:pharmacology; D014871:Water Microbiology
D021122:Protein Subunits; D012965:Sodium Chloride; D000101:Acetyl-CoA C-Acetyltransferase; D008274:Magnesium