Molecular cloning, characterization and purification of ornithine carbamoyltransferase from Mycobacterium bovis BCG.
A genomic library of Mycobacterium bovis BCG has been constructed by cloning DNA partially digested with Sau3A into the Escherichia coli expression vector pAS1. The gene coding for ornithine carbamoyl-transferase (EC.2.1.3.3; OTCase), hereafter referred to as argF, was isolated from the library by complementation of a double argF-argI mutant of E. coli and its sequence was determined. The translation initiation codon used, GTG, was identified by comparing the amino acid sequence deduced from the gene with the N-terminal sequence of the corresponding purified protein. On this basis, the M. bovis BCG OTCase monomer consists of 307 amino acid residues and displays about 44% identity with other OTCases, the most closely related homologue being the anabolic enzyme of Pseudomonas aeruginosa. The native enzyme has an estimated molecular mass of 110 kDa, suggesting a trimeric structure as is the case for most of the anabolic OTCases known from various organisms.
Timm
J
J
Service de Génétique Appliquée, Université Libre de Bruxelles, Nivelles Belgium.
Van Rompaey
I
I
Tricot
C
C
Massaer
M
M
Haeseleer
F
F
Fauconnier
A
A
Stalon
V
V
Bollen
A
A
Jacobs
P
P
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data; D009163:Mycobacterium bovis / Q000235:genetics*; D009954:Ornithine Carbamoyltransferase / Q000235:genetics* / Q000302:isolation & purification; D011401:Promoter Regions, Genetic; D016415:Sequence Alignment
D001426:Bacterial Proteins; D009954:Ornithine Carbamoyltransferase