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Kinetic analysis, structural studies and prediction of pKa values of Bacillus KR-8104 α-amylase: The determinants of pH-activity profile

Paper ID Volume ID Publish Year Pages File Format Full-Text
18732 42738 2007 9 PDF Available
Title
Kinetic analysis, structural studies and prediction of pKa values of Bacillus KR-8104 α-amylase: The determinants of pH-activity profile
Abstract

A native α-amylase capable of activity at low pH was isolated from Bacillus sp. KR-8104 (KRA) naturally occurred in rhizoplane zone of potato cultured soils. An investigation was set out to account some evidence for its specific properties. Kinetic parameters of KRA with respect to Bacillus licheniformis α-amylase (BLA) and B. amyloliquefaciens α-amylase (BAA) showed a shift in acidic limb for KRA pH-profile. After isolation of KRA gene and determination of its nucleotide and protein sequences, the three-dimensional structure of KRA was simulated based on BLA as template. Using PROPKA program the pKa values of titrable groups in KRA model together with other few members of this family were predicted either from simulated model or available structure at protein data bank. A correlation between catalytic residue pKa values and the optimal pH-activity profile was seen. Both our experimental data and prediction studies indicated a shift in pKa value of catalytic residues to acidic pH. Moreover alignment studies raised amino acid substitutions in KRA compare to BLA which may affect the KRA putative active site leading to the formation of an extra hydrogen bond between Glu261 and Arg229 (regarding to BLA numbering) and in turn shifts the pH-activity profile to lower amount. Also a common intermediate with characteristics of molten globular state was detected at a pH value lower than other members of α-amylase family using far UV circular dichroism, intrinsic and ANS fluorescence and quenching experiments.

Keywords
ANS, 1-anilino naphthalene-8-sulfonate; BAA, α-amylase from Bacillus amyloliquefaciens; BLA, α-amylase from Bacillus licheniformis; KRA, α-amylase from Bacillus sp. KR-8104; BStA, α-amylase from Bacillus stearothermophilus; CD, circular dichroismα-Amylase
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Kinetic analysis, structural studies and prediction of pKa values of Bacillus KR-8104 α-amylase: The determinants of pH-activity profile
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 41, Issue 3, 2 August 2007, Pages 337–345
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
Online Support
Any Questions? feel free to contact us