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Kinetic and thermodynamic properties of novel glucoamylase from Humicola sp.

Paper ID Volume ID Publish Year Pages File Format Full-Text
18635 42731 2007 7 PDF Available
Title
Kinetic and thermodynamic properties of novel glucoamylase from Humicola sp.
Abstract

Extra-cellular glucoamylase of Humicola sp. produced under submerged growth condition (10.44 U mg−1 protein) was purified to homogeneity level by using three-step purification procedure. Crude enzyme was subjected to ammonium sulphate precipitation, Hiload anion exchange chromatography and hydrophobic interaction column chromatography on FPLC purification system. The purified glucoamylase was monomeric in nature because native molecular mass on gel column chromatography and sub-unit mass on SDS-PAGE were the same, i.e. 72.8 kDa. Activation energy for soluble starch hydrolysis was 21.09 kJ mol−1, while temperature quotient (Q10) was 1.01. The enzyme was stable over a pH range of 3.5–5.9 and gave pH optimum of 4.7. Temperature optimum of the glucoamylase was 55 °C. Vmax for soluble starch hydrolysis was 56.63 U mg−1 protein and Km was 0.26 mg (soluble starch) ml−1. The turn over (kcat) was 69 s−1. The pKa1 and pKa2 of ionizable groups of active site controlling Vmax were 3.65 and 6.3, respectively. Thermodynamic parameters for soluble starch hydrolysis were as follows: ΔH* = 18.36 kJ mol−1, ΔG* = 69.06 kJ mol−1, ΔS* = −154.56 J mol−1 K−1, ΔGE−S*=−3.71 kJ mol−1 and ΔGE−T*=−26.41 kJ mol−1. Thermodynamic parameters (ΔH*, ΔG*, ΔS*) for irreversible inactivation of glucoamylase at different temperatures (45–55 °C) were also determined. Current report has novelty as it explained for the first time kinetics and thermodynamics of soluble starch hydrolysis and irreversible inactivation of glucoamylase from Humicola sp.

Keywords
Purification; Activation energy; Enthalpy; Entropy; Free energy
First Page Preview
Kinetic and thermodynamic properties of novel glucoamylase from Humicola sp.
Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 41, Issue 5, 1 October 2007, Pages 558–564
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering