A novel enzyme, citryl-CoA lyase, catalysing the second step of the citrate cleavage reaction in Hydrogenobacter thermophilus TK-6.
A novel enzyme catalysing citryl-CoA cleavage to acetyl-CoA and oxaloacetate was purified from Hydrogenobacter thermophilus TK-6, and designated citryl-CoA lyase (CCL). The citrate cleavage reaction in this organism proceeded by a unique set of two consecutive reactions: (i). citryl-CoA formation by citryl-CoA synthetase (CCS) and (ii). citryl-CoA cleavage by CCL. Purified CCL gave a single 30 kDa band in SDS-PAGE and gel filtration chromatography indicated that the native state of the enzyme exists as a trimer (alpha(3)). Citryl-CoA lyase showed low citrate synthase (CS) activity. Using an oligonucleotide probe, the corresponding gene was cloned and sequenced. The gene was expressed in Escherichia coli and recombinant CCL was also purified. The CCL protein sequence showed similarity to the C-terminal regions of ATP citrate lyase (ACL) and CS sequences in the database. By further sequence comparisons, the phylogenetic relationship between CCS, CCL, ACL and CS was investigated.
Aoshima
Miho
M
Department of Biotechnology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Yayoi 1-1-1, Bunkyo-ku, Tokyo 113-8657, Japan. aomihi@mail.ecc.u-tokyo.ac.jp
Ishii
Masaharu
M
Igarashi
Yasuo
Y
D000214:Acyl Coenzyme A / Q000378:metabolism*; D000595:Amino Acid Sequence; D001419:Bacteria / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D019343:Citric Acid / Q000378:metabolism*; D008969:Molecular Sequence Data; D007652:Oxo-Acid-Lyases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment
D000214:Acyl Coenzyme A; D001426:Bacterial Proteins; D011994:Recombinant Proteins; D019343:Citric Acid; C029890:citryl-coenzyme A; D007652:Oxo-Acid-Lyases; C110489:citryl-CoA lyase