Affiliation:
1. Korean Ocean Research & Development Institute, Ansan P.O. Box 29, Seoul 425-600, Korea
2. Division of Bioscience and Bioinformatics, Myongji University, Yongin 449-728, Korea
Abstract
ABSTRACT
Genomic analysis of a hyperthermophilic archaeon,
Thermococcus
sp. strain NA1, revealed the presence of a 1,068-bp open reading frame encoding a protein consisting of 356 amino acids with a calculated molecular mass of 39,714 Da (GenBank accession no. DQ144132). Sequence analysis showed that it was similar to the putative aminopeptidase P (APP) of
Thermococcus kodakaraensis
KOD1. Amino acid residues important for catalytic activity and the metal binding ligands conserved in bacterial, nematode, insect, and mammalian APPs were also conserved in the
Thermococcus
sp. strain NA1 APP. The archaeal APP, designated TNA1_APP (
Thermococcus
sp. strain NA1 APP), was cloned and expressed in
Escherichia coli
. The recombinant enzyme hydrolyzed the amino-terminal Xaa-Pro bond of Lys(
N
ε
-Abz)-Pro-Pro-pNA and the dipeptide Met-Pro (
K
m
, 0.96 mM), revealing its functional identity. Further enzyme characterization showed the enzyme to be a Co
2+
-, Mn
2+
-, or Zn
2+
-dependent metallopeptidase. Optimal APP activity with Met-Pro as the substrate occurred at pH 5 and a temperature of 100°C. The APP was thermostable, with a half-life of >100 min at 80°C. This study represents the first characterization of a hyperthermophilic archaeon APP.
Publisher
American Society for Microbiology
Subject
Ecology,Applied Microbiology and Biotechnology,Food Science,Biotechnology
Cited by
16 articles.
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