title: Genomic organisation, activity and distribution analysis of the microbial putrescine oxidase degradation pathway.
abstract: The catalytic action of putrescine specific amine oxidases acting in tandem with 4-aminobutyraldehyde dehydrogenase is explored as a degradative pathway in Rhodococcus opacus. By limiting the nitrogen source, increased catalytic activity was induced leading to a coordinated response in the oxidative deamination of putrescine to 4-aminobutyraldehyde and subsequent dehydrogenation to 4-aminobutyrate. Isolating the dehydrogenase by ion exchange chromatography and gel filtration revealed that the enzyme acts principally on linear aliphatic aldehydes possessing an amino moiety. Michaelis-Menten kinetic analysis delivered a Michaelis constant (K(M)=0.014 mM) and maximum rate (Vmax=11.2 μmol/min/mg) for the conversion of 4-aminobutyraldehyde to 4-aminobutyrate. The dehydrogenase identified by MALDI-TOF mass spectrometric analysis (E value=0.031, 23% coverage) belongs to a functionally related genomic cluster that includes the amine oxidase, suggesting their association in a directed cell response. Key regulatory, stress and transport encoding genes have been identified, along with candidate dehydrogenases and transaminases for the further conversion of 4-aminobutyrate to succinate. Genomic analysis has revealed highly similar metabolic gene clustering among members of Actinobacteria, providing insight into putrescine degradation notably among Micrococcaceae, Rhodococci and Corynebacterium by a pathway that was previously uncharacterised in bacteria.
authors.affiliation: Chemical Engineering, School of Engineering and Physical Science, Heriot-Watt University, Edinburgh EH 14 4AS, UK.
authors.forename: Alexander
authors.initials: A
authors.lastname: Foster
authors.forename: Nicole
authors.initials: N
authors.lastname: Barnes
authors.forename: Robert
authors.initials: R
authors.lastname: Speight
authors.forename: Mark A
authors.initials: MA
authors.lastname: Keane
mesh_terms: D000447:Aldehydes / Q000378:metabolism; D001711:Biotransformation; D003352:Corynebacterium / Q000235:genetics; D007700:Kinetics; D053858:Metabolic Networks and Pathways / Q000235:genetics*; D008835:Micrococcaceae / Q000235:genetics; D005810:Multigene Family; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D000587:Oxidoreductases Acting on CH-NH Group Donors / Q000235:genetics / Q000378:metabolism*; D011700:Putrescine / Q000378:metabolism*; D012240:Rhodococcus / Q000235:genetics* / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D019802:Succinic Acid / Q000378:metabolism; D005680:gamma-Aminobutyric Acid / Q000378:metabolism
chemical_list: D000447:Aldehydes; C006509:4-aminobutyraldehyde; D005680:gamma-Aminobutyric Acid; D019802:Succinic Acid; D010088:Oxidoreductases; C022488:putrescine oxidase; D000587:Oxidoreductases Acting on CH-NH Group Donors; D011700:Putrescine
keywords: Amine oxidase; Dehydrogenase; Phylogenetics; Putrescine degradation pathway