An arginine biosynthetic gene cluster, , of the extreme thermophilic bacterium HB27 was isolated by heterologous complementation of an acetylornithinase mutant. The recombinant plasmid (pTHM1) conferred ornithine acetyltransferase activity to the host, implying that uses the energetically more economic pathway for the deacetylation of acetylornithine. pTHM1 was, however, unable to complement an mutant and no acetylglutamate synthase activity could be detected in cells containing pTHM1. The -encoded enzyme is thus monofunctional and is unable to use acetyl-CoA to acetylate glutamate (contrary to the homologue). Alignment of several ornithine acetyltransferase amino acid sequences showed no obvious pattern that could account for this difference; however, the monofunctional enzymes proved to have shorter N-termini. Sequence analysis of the pTHM1 3.2 kb insert revealed the presence of the gene (encoding -acetylglutamate-5-semialdehyde dehydrogenase) upstream of the gene. Alignment of several -acetylglutamate-5-semialdehyde dehydrogenase amino acid sequences allowed identification of two strongly conserved putative motifs for cofactor binding: a putative FAD-binding site and a motif reminiscent of the NADPH-binding fingerprint. The relationship between the amino acid content of both enzymes and thermostability is discussed and an effect of the GC content bias is indicated. Transcription of both the and genes appeared to be vector-dependent. The -encoded enzyme activity was twofold repressed by arginine in the native host and was inhibited by ornithine. Both upstream of the gene and downstream of the gene an ORF with unknown function was found, indicating that the organization of the arginine biosynthetic genes in is new.


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