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Purification and characterization of a solvent stable aminopeptidase from Pseudomonas aeruginosa: Cloning and analysis of aminopeptidase gene conferring solvent stability
- Source :
-
Process Biochemistry . May2010, Vol. 45 Issue 5, p757-764. 8p. - Publication Year :
- 2010
-
Abstract
- Abstract: Aminopeptidase from a solvent tolerant strain Pseudomonas aeruginosa PseA was purified and studied for its biochemical and molecular characteristics. Ion-exchange chromatography resulted in 11.9-fold purification and 38% recovery of the 56kDa enzyme. The enzyme was found to be stable over a pH range of 6.0–8.0 and appreciably thermostable up to 70°C. PseA aminopeptidase exhibited K m of 3.02mM and V max of 6.71μmol/mg/min towards l-Leu-p-nitroanilide. Remarkable stability in both hydrophilic and hydrophobic solvents makes PseA aminopeptidase unique. Partial N-terminal sequence of enzyme showed exact match with probable aminopeptidase of P. aeruginosa PAO1, coded by gene pepB. Polymerase chain reaction amplified the 1611-bp open reading frame encoding a 57.51kDa, 536 amino acid PseA PepB polypeptide. The deduced PseA PepB protein sequence contained a 24-residue signal peptide (2.57kDa) followed by a 1.28kDa propeptide and a mature product of 500 residues. Search for conserved domain in PseA aminopeptidase explored its place in zinc-metallopeptidase family. Primary sequence analysis showed the hydrophobic inclination of the protein; and the 3D structure modeling elucidated the presence of a high content of hydrophobic residues on its surface probably imparting solvent stability to it. The enzyme might find potential applications in non-aqueous enzymology due to its marked thermostability and striking solvent stability. [Copyright &y& Elsevier]
Details
- Language :
- English
- ISSN :
- 13595113
- Volume :
- 45
- Issue :
- 5
- Database :
- Academic Search Index
- Journal :
- Process Biochemistry
- Publication Type :
- Academic Journal
- Accession number :
- 49110769
- Full Text :
- https://doi.org/10.1016/j.procbio.2010.01.017