title: gamma-Glutamylputrescine synthetase in the putrescine utilization pathway of Escherichia coli K-12.
abstract: Glutamate-putrescine ligase (gamma-glutamylputrescine synthetase, PuuA, EC 6.3.1.11) catalyzes the gamma-glutamylation of putrescine, the first step in a novel putrescine utilization pathway involving gamma-glutamylated intermediates, the Puu pathway, in Escherichia coli. In this report, the character and physiological importance of PuuA are described. Purified non-tagged PuuA catalyzed the ATP-dependent gamma-glutamylation of putrescine. The K(m) values for glutamate, ATP, and putrescine are 2.07, 2.35, and 44.6 mm, respectively. There are two putrescine utilization pathways in E. coli: the Puu pathway and the pathway without gamma-glutamylation. Gene deletion experiments of puuA, however, indicated that the Puu pathway was more critical in utilizing putrescine as a sole carbon or nitrogen source. The transcription of puuA was induced by putrescine and in a puuR-deleted strain. The amino acid sequences of PuuA and glutamine synthetase (GS) show high similarity. The molecular weights of the monomers of the two enzymes are quite similar, and PuuA exists as a dodecamer as does GS. Moreover the two amino acid residues of E. coli GS that are important for the metal-catalyzed oxidation of the enzyme molecule involved in protein turnover are conserved in PuuA, and it was experimentally shown that the corresponding amino acid residues in PuuA were involved in the metal-catalyzed oxidation similarly to GS. It is suggested that the intracellular concentration of putrescine is optimized by PuuA transcriptionally and posttranslationally and that excess putrescine is converted to a nutrient source by the Puu pathway.
authors.affiliation: Division of Integrated Life Science, Graduate School of Biostudies, Kyoto University, Kyoto, Japan.
authors.forename: Shin
authors.initials: S
authors.lastname: Kurihara
authors.forename: Shinpei
authors.initials: S
authors.lastname: Oda
authors.forename: Yuichi
authors.initials: Y
authors.lastname: Tsuboi
authors.forename: Hyeon Guk
authors.initials: HG
authors.lastname: Kim
authors.forename: Mayu
authors.initials: M
authors.lastname: Oshida
authors.forename: Hidehiko
authors.initials: H
authors.lastname: Kumagai
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Suzuki
mesh_terms: D000595:Amino Acid Sequence; D000641:Ammonia / Q000378:metabolism; D020134:Catalytic Domain; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D008025:Ligases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D005810:Multigene Family / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D011700:Putrescine / Q000378:metabolism*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D000637:Transaminases / Q000378:metabolism; D014158:Transcription, Genetic / Q000235:genetics
chemical_list: D029968:Escherichia coli Proteins; D000641:Ammonia; D000637:Transaminases; C507750:YgjG protein, E coli; D008025:Ligases; C530996:gamma-glutamylputrescine synthetase, E coli; D011700:Putrescine