Kinetic and Thermodynamic Characterization of Glucoamylase from Colletotrichum sp. KCP1

Abstract

Extracellular glucoamylase of Colletotrichum sp. KCP1 produced through solid state fermentation was purified by two steps purification process comprising ammonium sulphate precipitation followed by gel permeation chromatography (GPC). The Recovery of glucoamylase after GPC was 50.40 % with 19.3-fold increase in specific activity. The molecular weight of enzyme was found to be 162.18 kDa by native-PAGE and was dimeric protein of two sub-units with molecular weight of 94.62 and 67.60 kDa as determined by SDS-PAGE. Activation energy for starch hydrolysis was 26.45 kJ mol -1 while temperature quotient (Q 10 ) was found to be 1.9. The enzyme was found to be stable over wide pH range and thermally stable at 40-50°C up to 120 min while exhibited maximum activity at 50°C with pH 5.0. The pKa 1 and pKa 2 of ionisable groups of active site controlling V max were 3.5 and 6.8, respectively. V max , K m and K cat for starch hydrolysis were found to be 58.82 U ml -1 , 1.17 mg (starch) ml -1 and 449 s -1 , respectively. Activation energy for irreversible inactivation (E a(d) ) of glucoamylase was 74.85 kJ mol -1 . Thermodynamic parameters of irreversible inactivation of glucoamylase and starch hydrolysis were also determined.

Purification and characterization of a novel α-glucosidase from an Antarctic yeast Dioszegia fristingensis isolate

“…Comparatively, this is much lower than seen for a glucoamylase from Colletotrichum sp. KCP1, which showed a good stability at 30-50 o C with decimal reduction time values higher than 17 h within that temperature interval [51]. The decimal reduction time value of the D. fristingensis α-glucosidase was approximately 70 h. However, this is only a preliminary characterization and additional thermodynamic studies must be performed to understand the enzyme thermal behaviour and its activity-stability trade-off [47].…”

Dietary polysaccharides: fermentation potentials of a primitive gut ecosystem

“…It can be postulated that early gut ecosystems were at least partially dependent on microbes capable of utilizing the 'easiest substrates' first. Rather, more easily hydrolyzed polysaccharides such as starch [activation energy approximates 26 kJ per mol (Prajapati et al, 2014)] would have been more likely candidates as sources of polysaccharide-derived energy. Thus, it seems unlikely that the use of difficult to degrade structural polysaccharides such as cellulose, a polymer with extensive hydrogen bonding to ensure stability [activation energy ranges from 95 to 130 kJ per mol (Kunov-Kruse et al, 2013;Sørensen et al, 2015)] could have been prioritized as the digestive system of the earthworm was evolving.…”

“…Thus, it seems unlikely that the use of difficult to degrade structural polysaccharides such as cellulose, a polymer with extensive hydrogen bonding to ensure stability [activation energy ranges from 95 to 130 kJ per mol (Kunov-Kruse et al, 2013;Sørensen et al, 2015)] could have been prioritized as the digestive system of the earthworm was evolving. Rather, more easily hydrolyzed polysaccharides such as starch [activation energy approximates 26 kJ per mol (Prajapati et al, 2014)] would have been more likely candidates as sources of polysaccharide-derived energy. Indeed, even ruminants that have highly specialized cellulosedegrading microbiota and utilize rumination to maximize breakdown of cellulolytic fibres are unable to completely digest all of the cellulose they ingest (Russell et al, 2009).…”

Evaluation of Catalytic Efficiency of Coriolopsis caperata DN Laccase to Decolorize and Detoxify RBBR Dye

“…Influence of temperature towards decolorization of dyes was studied at optimum pH at different temperature ranges (10-50°C). Effect of temperature on the rate of dye decolorization was expressed in terms of temperature quotient (Q 10 ) value which is a measure of the temperature sensitivity of an enzymatic reaction rate by 10°C [13].…”