Thermodynamics and kinetic properties of halostable endoglucanase from Aspergillus fumigatus ABK9

Abstract

An endoglucanase from Aspergillus fumigatus ABK9 was purified from the culture extract of solid-state fermentation and its some characteristics were evaluated. The molecular weight of the purified enzyme (56.3 kDa) was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, zymogram analysis and confirmed by MALDI-TOF mass spectrometry. The enzyme was active optimally at 50 °C, pH 5.0 and stable over a broad range of pH (4.0-7.0) and NaCl concentration of 0-3.0 M. The pKa1 and pKa2 of the ionizable groups of the active sites were 2.94 and 6.53, respectively. The apparent Km , Vmax , and Kcat values for carboxymethyl cellulose were 6.7 mg ml(-1), 775.4 µmol min(-1) , and 42.84 × 10(4) s(-1), respectively. Thermostability of the enzyme was evidenced by the high activation energy (91.45 kJ mol(-1)), large enthalpy for activation of denaturation (88.77 kJ mol(-1)), longer half-life (T1/2) (433 min at 50 °C), higher melting temperature (Tm ) (73.5 °C), and Q10 (1.3) values. All the characteristics favors its suitability as halotolerant and thermostable enzyme during bioprocessing of lignocellulosic materials.

Proteomic analysis of Streptomyces sp. ssr‐198 grown on paddy straw

“…The culture of Streptomyces sp. ssr-198 was grown in an optimized sterilized medium (untreated paddy straw 18.39 g, peptone 1.0 g, K 2 HPO 4 5.0 g, (NH 4 )SO 4 . For inoculum preparation, the 24 h old cultures grown in nutrient broth were centrifuged and re-suspended in normal saline to achieve an OD of 0.8.…”

“…and other enzymes from different microorganisms to produce high titer of a synergistically acting enzyme cocktail. Besides T. reesei , many other filamentous fungi like T. viride , Humicola insolens , Penicillium verruculosum , A. japonicus , A. fumigatus , Acremonium cellulolyticus are also known to be good producers of different cellulolytic enzymes .…”

Cloning and expression of an endoglucanase gene from the thermotolerant fungus Aspergillus fumigatus DBiNU-1 in Kluyveromyces lactis

“…The results of the current study were in good agreement with those reported for most of the endoglucanase from Aspergillus strains, such as A. terreus DSM 826, 42 A. awamori VTCC-F099, 38 A. usamii E001 7 and A. fumigatus ABK9. 20 However, a high thermal stability (70 °C) was also reported for endoglucanases from T. aurantiacus 43 and A. fumigatus MKU1. 3 The differences in thermal stability indicated the different molecular properties of the enzymes, including the bonding that stabilizes in the structures and the conformations among the various species.…”

“…The predicted molecular weight of the endoglucanase from A. fumigatus DBiNU-1, as confirmed by SDS-PAGE analysis, was approximately 48 kDa, which was distinct from other endoglucanases from other Aspergillus strains, e.g., A. terreus M11 (25 kDa), 37 A. awamori VTCC-F099 (32 kDa), 38 A. fumigatus KMU1 (35 kDa), 3 A. oryzae AG1 (45 kDa), 39 A. usamii E001 (24 kDa), 7 and A. fumigatus ABK9 (56 kDa). 20 Many glycoside hydrolases such as GH7, GH12, GH16 and AA9 have a modular structure that consists of a catalytic domain and a substrate-binding domain. 40 The Swiss-PdbViewer ( http://www/expasy.org/spdbv ) was used to predict the three-dimensional structure of the putative amino acid sequence of the endoglucanase from A. fumigatus DBiNU-1 (Additional information section, Fig.…”

“…Most recently, the endoglucanase gene belonging to the family AA9 (formerly known as GH61) from the alkali-tolerant A. fumigatus MKU1, which is isolated from paper and pulp industry wastes, has also been cloned and sequenced. 3 Das et al 20 reported the kinetic properties of the purified halostable endoglucanase from a culture extract of A. fumigatus ABK9, and the authors demonstrated that the calculated molecular weight of the enzyme was approximately 56.3 kDa. The enzyme displayed its maximum activity at a temperature of 50 °C and a pH value of 5.0.…”