Identification of the putrescine biosynthetic genes in Pseudomonas aeruginosa and characterization of agmatine deiminase and N-carbamoylputrescine amidohydrolase of the arginine decarboxylase pathway.
Putrescine can be synthesized either directly from ornithine by ornithine decarboxylase (ODC; the speC product) or indirectly from arginine via arginine decarboxylase (ADC; the speA product). The authors identified the speA and speC genes in Pseudomonas aeruginosa PAO1. The activities of the two decarboxylases were similar and each enzyme alone appeared to direct sufficient formation of the polyamine for normal growth. A mutant defective in both speA and speC was a putrescine auxotroph. In this strain, agmatine deiminase (the aguA product) and N-carbamoylputrescine amidohydrolase (the aguB product), which were initially identified as the catabolic enzymes of agmatine, biosynthetically convert agmatine to putrescine in the ADC pathway: a double mutant of aguAB and speC was a putrescine auxotroph. AguA was purified as a homodimer of 43 kDa subunits and AguB as a homohexamer of 33 kDa subunits. AguA specifically deiminated agmatine with K(m) and K(cat) values of 0.6 mM and 4.2 s(-1), respectively. AguB was specific to N-carbamoylputrescine and the K(m) and K(cat) values of the enzyme for the substrate were 0.5 mM and 3.3 s(-1), respectively. Whereas AguA has no structural relationship to any known C-N hydrolases, AguB is a protein of the nitrilase family that performs thiol-assisted catalysis. Inhibition by SH reagents and the conserved cysteine residue in AguA and its homologues suggested that this enzyme is also involved in thiol-mediated catalysis.
Nakada
Yuji
Y
Division of Applied Microbiology, National Food Research Institute, Kannondai 2-1-12, Tsukuba Ibaraki 305-8642, Japan.
Itoh
Yoshifumi
Y
Division of Applied Microbiology, National Food Research Institute, Kannondai 2-1-12, Tsukuba Ibaraki 305-8642, Japan.
D000595:Amino Acid Sequence; D002262:Carboxy-Lyases / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D006867:Hydrolases / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D009955:Ornithine Decarboxylase / Q000235:genetics / Q000378:metabolism; D011550:Pseudomonas aeruginosa / Q000201:enzymology* / Q000235:genetics; D011700:Putrescine / Q000096:biosynthesis*; D014512:Ureohydrolases / Q000235:genetics / Q000378:metabolism*
D006867:Hydrolases; C027237:N-carbamoylputrescine hydrolase; D014512:Ureohydrolases; C019626:agmatine deiminase; D002262:Carboxy-Lyases; D009955:Ornithine Decarboxylase; C019722:arginine decarboxylase; D011700:Putrescine