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Purification and characterization of a solvent stable aminopeptidase from Pseudomonas aeruginosa: Cloning and analysis of aminopeptidase gene conferring solvent stability

Paper ID Volume ID Publish Year Pages File Format Full-Text
35682 45101 2010 8 PDF Available
Title
Purification and characterization of a solvent stable aminopeptidase from Pseudomonas aeruginosa: Cloning and analysis of aminopeptidase gene conferring solvent stability
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 56 kDa 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 Km of 3.02 mM and Vmax of 6.71 μmol/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 57.51 kDa, 536 amino acid PseA PepB polypeptide. The deduced PseA PepB protein sequence contained a 24-residue signal peptide (2.57 kDa) followed by a 1.28 kDa 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.

Keywords
Pseudomonas aeruginosa; Aminopeptidase; Ion-exchange chromatography; Metallopeptidase; Solvent tolerant
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Purification and characterization of a solvent stable aminopeptidase from Pseudomonas aeruginosa: Cloning and analysis of aminopeptidase gene conferring solvent stability
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Publisher
Database: Elsevier - ScienceDirect
Journal: Process Biochemistry - Volume 45, Issue 5, May 2010, Pages 757–764
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
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Any Questions? feel free to contact us