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Purification and characterization of a novel pectinase from Acrophialophora nainiana with emphasis on its physicochemical properties

Paper ID Volume ID Publish Year Pages File Format Full-Text
25776 43599 2006 10 PDF Available
Title
Purification and characterization of a novel pectinase from Acrophialophora nainiana with emphasis on its physicochemical properties
Abstract

An extracellular pectinase (PECI) was purified to apparent homogeneity from liquid state cultures of the thermophilic fungus Acrophialophora nainiana by ultrafiltration and a combination of gel filtration and ion-exchange chromatographic procedures. The molecular masses of PECI were 35,500 and 30,749 Da, as determined by SDS-PAGE and mass spectrometry, respectively. It was more active at 60 °C and pH 8.0 and showed high stability at 50 °C with half-life of 7 days. However at 60 and 70 °C, PECI was much less stable with half lives of approximately 20 and 3 min, respectively. The thermostability of purified PECI was also investigated by fluorescence and circular dichroism spectroscopy. Fluorescence revealed that the unfolding transition region was observed between 45 and 70 °C. A major decrease in the stability was found at 70 °C. Circular dichroism measurements at pH between 5.0 and 9.0 showed a transition temperature (Tm) range of 50–55°. The thermodynamic analysis of these results showed that EPGI is thermal stable protein exhibiting maximum stability (ΔG25) of 22.65 and 19.19 kcal/mol at pH 8.0 and 9.0, respectively. The apparent Km value on pectin from citrus fruits was 4.22 mg ml−1. PECI exhibited no detectable activity of pectin methylesterase, endo-polygalacturonase, mannanase, xylanase and cellulase. However, it showed exo-polygalacturonase and pectin lyase activities. The presence of carbohydrate was detected in the pure PECI. It was activated by l-tryptophan, DEPC, DTT, DTNB, DTP, l-cystein and β-mercaptoethanol and inhibited by NBS, Fe2+, Cu2+, Zn2+, Mn2+, Al3+ and Ca2+. The enzyme showed homology with a pectin lyases from Xanthomonas campestris and Bacillus licheniformis.

Keywords
Acrophialophora nainiana; Pectin; Pectinase; Thermostability
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Purification and characterization of a novel pectinase from Acrophialophora nainiana with emphasis on its physicochemical properties
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 123, Issue 1, 3 May 2006, Pages 33–42
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