title: The first agmatine/cadaverine aminopropyl transferase: biochemical and structural characterization of an enzyme involved in polyamine biosynthesis in the hyperthermophilic archaeon Pyrococcus furiosus.
abstract: We report here the characterization of the first agmatine/cadaverine aminopropyl transferase (ACAPT), the enzyme responsible for polyamine biosynthesis from an archaeon. The gene PF0127 encoding ACAPT in the hyperthermophile Pyrococcus furiosus was cloned and expressed in Escherichia coli, and the recombinant protein was purified to homogeneity. P. furiosus ACAPT is a homodimer of 65 kDa. The broad substrate specificity of the enzyme toward the amine acceptors is unique, as agmatine, 1,3-diaminopropane, putrescine, cadaverine, and sym-nor-spermidine all serve as substrates. While maximal catalytic activity was observed with cadaverine, agmatine was the preferred substrate on the basis of the k(cat)/K(m) value. P. furiosus ACAPT is thermoactive and thermostable with an apparent melting temperature of 108 degrees C that increases to 112 degrees C in the presence of cadaverine. Limited proteolysis indicated that the only proteolytic cleavage site is localized in the C-terminal region and that the C-terminal peptide is not necessary for the integrity of the active site. The crystal structure of the enzyme determined to 1.8-A resolution confirmed its dimeric nature and provided insight into the proteolytic analyses as well as into mechanisms of thermal stability. Analysis of the polyamine content of P. furiosus showed that spermidine, cadaverine, and sym-nor-spermidine are the major components, with small amounts of sym-nor-spermine and N-(3-aminopropyl)cadaverine (APC). This is the first report in Archaea of an unusual polyamine APC that is proposed to play a role in stress adaptation.
authors.affiliation: Dipartimento di Biochimica e Biofisica, F. Cedrangolo, Seconda Università degli Studi di Napoli, Via Costantinopoli 16, 80138 Naples, Italy. giovanna.cacciapuoti@unina2.it
authors.forename: Giovanna
authors.initials: G
authors.lastname: Cacciapuoti
authors.forename: Marina
authors.initials: M
authors.lastname: Porcelli
authors.forename: Maria Angela
authors.initials: MA
authors.lastname: Moretti
authors.forename: Francesca
authors.initials: F
authors.lastname: Sorrentino
authors.forename: Luigi
authors.initials: L
authors.lastname: Concilio
authors.forename: Vincenzo
authors.initials: V
authors.lastname: Zappia
authors.forename: Zhi-Jie
authors.initials: ZJ
authors.lastname: Liu
authors.forename: Wolfram
authors.initials: W
authors.lastname: Tempel
authors.forename: Florian
authors.initials: F
authors.lastname: Schubot
authors.forename: John P
authors.initials: JP
authors.lastname: Rose
authors.forename: Bi-Cheng
authors.initials: BC
authors.lastname: Wang
authors.forename: Phillip S
authors.initials: PS
authors.lastname: Brereton
authors.forename: Francis E
authors.initials: FE
authors.lastname: Jenney
authors.forename: Michael W W
authors.initials: MW
authors.lastname: Adams
mesh_terms: D000376:Agmatine / Q000378:metabolism; D019883:Alkyl and Aryl Transferases / Q000096:biosynthesis; D001105:Archaea; D019843:Archaeal Proteins / Q000096:biosynthesis; D002103:Cadaverine / Q000378:metabolism; D019766:Genes, Archaeal; D006358:Hot Temperature; D011073:Polyamines / Q000378:metabolism*; D020027:Pyrococcus furiosus / Q000201:enzymology* / Q000235:genetics / Q000502:physiology*; D011436:Spermidine Synthase / Q000737:chemistry / Q000378:metabolism
chemical_list: D019843:Archaeal Proteins; D011073:Polyamines; D000376:Agmatine; D019883:Alkyl and Aryl Transferases; C524065:agmatine-cadaverine aminopropyl transferase, Pyrococcus furiosus; D011436:Spermidine Synthase; D002103:Cadaverine