title: Amine-synthesizing enzyme N-substituted formamide deformylase: screening, purification, characterization, and gene cloning.
abstract: N-substituted formamide was produced through the hydration of an isonitrile by isonitrile hydratase in the isonitrile metabolism. The former compound was further degraded by a microorganism, strain F164, which was isolated from soil through an acclimatization culture. The N-substituted formamide-degrading microorganism was identified as Arthrobacter pascens. The microbial degradation was found to proceed through an enzymatic reaction, the N-substituted formamide being hydrolyzed to yield the corresponding amine and formate. The enzyme, designated as N-substituted formamide deformylase (NfdA), was purified and characterized. The native enzyme had a molecular mass of approximately 61 kDa and consisted of two identical subunits. It stoichiometrically catalyzed the hydrolysis of N-benzylformamide (an N-substituted formamide) to benzylamine and formate. Of all of the N-substituted formamides tested, N-benzylformamide was the most suitable substrate for the enzyme. However, no amides were accepted as substrates. The gene (nfdA) encoding this enzyme was also cloned. The deduced amino acid sequence of nfdA exhibited the highest overall sequence identity (28%) with those of regulatory proteins among known proteins. Only the N-terminal region (residues 58-72) of NfdA also showed significant sequence identity (27-73%) to that of each member of the amidohydrolase superfamily, although there was no similarity in the overall sequence except in the above limited region.
authors.affiliation: Institute of Applied Biochemistry and Graduate School of Life and Environmental Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8572, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Fukatsu
authors.forename: Yoshiteru
authors.initials: Y
authors.lastname: Hashimoto
authors.forename: Masahiko
authors.initials: M
authors.lastname: Goda
authors.forename: Hiroki
authors.initials: H
authors.lastname: Higashibata
authors.forename: Michihiko
authors.initials: M
authors.lastname: Kobayashi
mesh_terms: D000581:Amidohydrolases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D000588:Amines / Q000138:chemical synthesis*; D000595:Amino Acid Sequence; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular / Q000379:methods; D017124:Conserved Sequence; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
chemical_list: D000588:Amines; D011994:Recombinant Proteins; D000581:Amidohydrolases; C493455:N-substituted formamide deformylase