title: Detection and characterization of a thermophilic biotin biosynthetic enzyme, 7-keto-8-aminopelargonic acid synthase, from various thermophiles.
abstract: By detailed BLAST searches of the genome database of various thermophiles, five ORFs with similarity to the bioF gene, which encodes 7-keto-8-aminopelargonic acid synthase (BioF) involved in biotin biosynthesis, of Escherichia coli were found: AqbioF, CltbioF, GkbioF, SytbioF, and TsebioF, from Aquifex aeolicus VF5, Clostridium thermocellum ATCC27405, Geobacillus kaustophilus JCM12893, Symbiobacterium thermophilum IAM14863, and Thermosynechococcus elongatus BP-1 respectively. The five purified recombinant bioF gene products, which were overexpressed in E. coli, had the enzyme activity of BioF. The optimum temperature range and thermostability of five BioFs, AqBioF, CltBioF, GkBioF, SytBioF, and TseBioF, were higher than those of E. coli BioF. In particular, AqBioF was found to show the highest thermostability of the α-oxoamine synthase family enzymes reported to date. Substrate specificity experiments revealed that SytBioF was also able to catalyze the reaction of 2-amino-3-ketobutyrate CoA ligase, a member of the α-oxoamine synthase family, and that it used acetyl-CoA and glycine as substrates, like the TTHA1582 protein of Thermus thermophilus. The other purified BioFs, AqBioF and GkBioF, did not show any activity with acyl-CoAs and amino acids other than pimeloyl-CoA and L-alanine as substrates.
authors.affiliation: Department of Biotechnology, Tottori University, Tottori, Japan.
authors.forename: Takaaki
authors.initials: T
authors.lastname: Kubota
authors.forename: Yoshikazu
authors.initials: Y
authors.lastname: Izumi
mesh_terms: D000123:Acetyltransferases / Q000378:metabolism; D000214:Acyl Coenzyme A / Q000378:metabolism; D000217:Acyltransferases / Q000235:genetics / Q000378:metabolism*; D000409:Alanine / Q000378:metabolism; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001710:Biotin / Q000096:biosynthesis* / Q000235:genetics; D003001:Cloning, Molecular; D004795:Enzyme Stability; D004926:Escherichia coli; D015870:Gene Expression; D005998:Glycine / Q000378:metabolism; D006094:Gram-Positive Bacteria / Q000201:enzymology* / Q000235:genetics; D006358:Hot Temperature; D007651:Keto Acids / Q000378:metabolism; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013379:Substrate Specificity
chemical_list: D000214:Acyl Coenzyme A; D000596:Amino Acids; D001426:Bacterial Proteins; D007651:Keto Acids; D011994:Recombinant Proteins; C019527:pimeloyl-coenzyme A; C079776:2-amino-3-ketobutyrate; D001710:Biotin; D000217:Acyltransferases; D000123:Acetyltransferases; C020679:glycine acetyltransferase; C021929:8-amino-7-oxononanoate synthase; D000409:Alanine; D005998:Glycine