title: Molecular characterization of a novel N-acetyltransferase from Chryseobacterium sp.
abstract: N-Acetyltransferase from Chryseobacterium sp. strain 5-3B is an acetyl coenzyme A (acetyl-CoA)-dependent enzyme that catalyzes the enantioselective transfer of an acetyl group from acetyl-CoA to the amino group of l-2-phenylglycine to produce (2S)-2-acetylamino-2-phenylacetic acid. We purified the enzyme from strain 5-3B and deduced the N-terminal amino acid sequence. The gene, designated natA, was cloned with two other hypothetical protein genes; the three genes probably form a 2.5-kb operon. The deduced amino acid sequence of NatA showed high levels of identity to sequences of putative N-acetyltransferases of Chryseobacterium spp. but not to other known arylamine and arylalkylamine N-acetyltransferases. Phylogenetic analysis indicated that NatA forms a distinct lineage from known N-acetyltransferases. We heterologously expressed recombinant NatA (rNatA) in Escherichia coli and purified it. rNatA showed high activity for l-2-phenylglycine and its chloro- and hydroxyl-derivatives. The Km and Vmax values for l-2-phenylglycine were 0.145 ± 0.026 mM and 43.6 ± 2.39 μmol · min(-1) · mg protein(-1), respectively. The enzyme showed low activity for 5-aminosalicylic acid and 5-hydroxytryptamine, which are reported as good substrates of a known arylamine N-acetyltransferase and an arylalkylamine N-acetyltransferase. rNatA had a comparatively broad acyl donor specificity, transferring acyl groups to l-2-phenylglycine and producing the corresponding 2-acetylamino-2-phenylacetic acids (relative activity with acetyl donors acetyl-CoA, propanoyl-CoA, butanoyl-CoA, pentanoyl-CoA, and hexanoyl-CoA, 100:108:122:10:<1).
authors.affiliation: Department of Agrobioscience, Graduate School of Agricultural Science, Kobe University, Kobe, Japan.
authors.forename: Shinji
authors.initials: S
authors.lastname: Takenaka
authors.forename: Kenji
authors.initials: K
authors.lastname: Yoshida
authors.forename: Kosei
authors.initials: K
authors.lastname: Tanaka
authors.forename: Ken-Ichi
authors.initials: K
authors.lastname: Yoshida
mesh_terms: D000105:Acetyl Coenzyme A / Q000378:metabolism*; D000123:Acetyltransferases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D045247:Chryseobacterium / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D005998:Glycine / Q000031:analogs & derivatives* / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D010648:Phenylacetates / Q000378:metabolism*; D010802:Phylogeny; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
chemical_list: D004269:DNA, Bacterial; D010648:Phenylacetates; D011994:Recombinant Proteins; C008852:2-phenylglycine; D000105:Acetyl Coenzyme A; D000123:Acetyltransferases; D005998:Glycine