Evolution in the amidohydrolase superfamily: Substrate-assisted gain of function in the E183K mutant of a phosphotriesterase-like metal-carboxylesterase (Articolo in rivista)

Type
Label
  • Evolution in the amidohydrolase superfamily: Substrate-assisted gain of function in the E183K mutant of a phosphotriesterase-like metal-carboxylesterase (Articolo in rivista) (literal)
Anno
  • 2009-01-01T00:00:00+01:00 (literal)
Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#doi
  • 10.1021/bi801932x (literal)
Alternative label
  • Mandrich L.; Manco G. (2009)
    Evolution in the amidohydrolase superfamily: Substrate-assisted gain of function in the E183K mutant of a phosphotriesterase-like metal-carboxylesterase
    in Biochemistry (Easton)
    (literal)
Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#autori
  • Mandrich L.; Manco G. (literal)
Pagina inizio
  • 5602 (literal)
Pagina fine
  • 5612 (literal)
Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#numeroVolume
  • 48 (literal)
Rivista
Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#numeroFascicolo
  • 24 (literal)
Note
  • ISI Web of Science (WOS) (literal)
Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#affiliazioni
  • Institute of Protein Biochemistry, National Research Council, Via Pietro Castellino 111, 80131, Naples, Italy (literal)
Titolo
  • Evolution in the amidohydrolase superfamily: Substrate-assisted gain of function in the E183K mutant of a phosphotriesterase-like metal-carboxylesterase (literal)
Abstract
  • The recent specialization for utilization of pesticides reported for Pseudomonas diminuta phosphotriesterase (pPTE), strongly suggests that this activity evolved from an enzyme endowed with promiscuous phosphotriesterase activity. Such putative \"generalist\" enzyme was recently proposed to be a member of the new phoshotriesterase-like lactonase family (PLL). The promiscuous carboxylesterase and phosphodiesterase activities detected in pPTE and PLLs, in turn paved the way for the prediction of the existence in nature of PTE-like enzymes with predominant carboxylesterase or phosphodiesterase activities. An \"in silico\" analysis of the related Mesorhizobium loti ORF MLL7664 and the biochemical characterization demonstrated its prominent carboxylesterase and low phosphotriesterase specificity. Based on sequence similarity with the Phosphotriesterase Homology Protein from E. coli and the carboxylesterase activity, we called it phosphotriesterase-like carboxylesterase (MloPLC). The carboxylesterase activity is strictly dependent on divalent cations, and as such MloPLC is the first phosphotriesterase-like metal-carboxylesterase characterized to date. In related enzymes of the amidohydrolase superfamily either a glutamate or a carboxylated lysine substitutes for MloPLC glutamate 183, and the residue appears invariantly involved in maintaining the structural integrity of the binuclear metal center. Accordingly, we changed Glu-183 to lysine or glutamine. All the tested activities were completely abolished in the E183Q mutant, while only a residual phosphotriesterase activity could be detected in the E183K mutant. Surprisingly, in the latter mutant a parallel 650-fold specificity increase on bis-p-nitrophenyl-phosphate (BpNP-P) was observed, turning MloPLC from a carboxylesterase into a phosphodiesterase. Chemical, structural and kinetic data strongly suggested that K183 is not carboxylated, and that the gain of the new function is assisted by the substrate. (literal)
Prodotto di
Autore CNR
Insieme di parole chiave

Incoming links:


Autore CNR di
Prodotto
Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#rivistaDi
Insieme di parole chiave di
data.CNR.it