[Novel L-amino acid ligases catalyzing oligopeptide synthesis].
L-Amino acid ligase (EC 6.3.2.28) is a microbial enzyme catalyzing formation of an alpha-peptide bond from unprotected L-amino acids in an ATP-dependent manner. The YwfE protein from Bacillus subtilis 168 was the first reported L-amino acid ligase, and it synthesizes various dipeptides. Thereafter, several L-amino acid ligases were newly obtained by in silico analysis using the ATP-grasp motif. But these L-amino acid ligases synthesize only dipeptide and no longer peptide. A novel L-amino acid ligase capable of catalyzing oligopeptide synthesis is required to increase the variety of peptides. We have previously found a new member of L-amino acid ligase, RizA, from B. subtilis NBRC3134, a microorganism that produces the peptide-antibiotic rhizocticin. We newly found that a gene at approximately 9 kbp upstream of rizA encoded a novel L-amino acid ligase RizB. Recombinant RizB synthesized homo-oligomers of branched-chain amino acids consisting of 2 to 5 amino acids, and also synthesized various heteropeptides. RizB is the first reported L-amino acid ligase that catalyzes oligopeptide synthesis. In addition, we obtained L-amino acid ligases showing oligopeptide synthesis activities by in silico analysis using BLAST, which is a set of similarity search programs. These L-amino acid ligases showed low similarity in amino acid sequence, but commonly used branched-chain amino acids, such as RizB, as substrates. Furthermore, the spr0969 protein of Streptococcus pneumoniae synthesized longer peptides than those synthesized by RizB, and the BAD_1200 protein of Bifidobacteria adolescentis showed higher activity toward aromatic amino acids than toward branched-chain ones.
Department of Applied Chemistry, School of Advanced Science and Engineering, Waseda University, Shinjuku-ku, Japan. kkino.@waseda,jp
Kuniki
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Kino
D000596:Amino Acids; D000597:Amino Acids, Branched-Chain / Q000378:metabolism; D001412:Bacillus subtilis / Q000201:enzymology / Q000378:metabolism; D055162:Biocatalysis; D057166:High-Throughput Screening Assays; D008025:Ligases / Q000235:genetics / Q000502:physiology*; D009842:Oligopeptides / Q000096:biosynthesis*; D009943:Organophosphorus Compounds; D011994:Recombinant Proteins
D000596:Amino Acids; D000597:Amino Acids, Branched-Chain; D009842:Oligopeptides; D009943:Organophosphorus Compounds; D011994:Recombinant Proteins; C063847:rhizocticin A; D008025:Ligases