Characterization of bacterial NMN deamidase as a Ser/Lys hydrolase expands diversity of serine amidohydrolases

FEBS Lett. 2014 Mar 18;588(6):1016-23. doi: 10.1016/j.febslet.2014.01.063. Epub 2014 Feb 11.

Abstract

NMN deamidase (PncC) is a bacterial enzyme involved in NAD biosynthesis. We have previously demonstrated that PncC is structurally distinct from other known amidohydrolases. Here, we extended PncC characterization by mutating all potential catalytic residues and assessing their individual roles in catalysis through kinetic analyses. Inspection of these residues' spatial arrangement in the active site, allowed us to conclude that PncC is a serine-amidohydrolase, employing a Ser/Lys dyad for catalysis. Analysis of the PncC structure in complex with a modeled NMN substrate supported our conclusion, and enabled us to propose the catalytic mechanism.

Keywords: Amidohydrolase; Catalytic dyad; NMN deamidase; Pyridine nucleotide; Site-directed mutagenesis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amidohydrolases / chemistry*
  • Amidohydrolases / genetics
  • Amino Acid Sequence
  • Amino Acid Substitution
  • Apoenzymes / chemistry
  • Apoenzymes / genetics
  • Catalytic Domain
  • Conserved Sequence
  • Enzyme Stability
  • Escherichia coli Proteins / chemistry*
  • Escherichia coli Proteins / genetics
  • Kinetics
  • Models, Molecular
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Nicotinamide Mononucleotide / chemistry
  • Protein Structure, Secondary
  • Sequence Homology, Amino Acid

Substances

  • Apoenzymes
  • Escherichia coli Proteins
  • Nicotinamide Mononucleotide
  • Amidohydrolases
  • nicotinamidenucleotide amidase