Cellulase complex of the fungus Chrysosporium lucknowense: isolation and characterization of endoglucanases and cellobiohydrolases

Biochemistry (Mosc). 2004 May;69(5):542-51. doi: 10.1023/b:biry.0000029853.34093.13.

Abstract

Using different chromatographic techniques, eight cellulolytic enzymes were isolated from the culture broth of a mutant strain of Chrysosporium lucknowense: six endoglucanases (EG: 25 kD, pI 4.0; 28 kD, pI 5.7; 44 kD, pI 6.0; 47 kD, pI 5.7; 51 kD, pI 4.8; 60 kD, pI 3.7) and two cellobiohydrolases (CBH I, 65 kD, pI 4.5; CBH II, 42 kD, pI 4.2). Some of the isolated cellulases were classified into known families of glycoside hydrolases: Cel6A (CBH II), Cel7A (CBH I), Cel12A (EG28), Cel45A (EG25). It was shown that EG44 and EG51 are two different forms of one enzyme. EG44 seems to be a catalytic module of an intact EG51 without a cellulose-binding module. All the enzymes had pH optimum of activity in the acidic range (at pH 4.5-6.0), whereas EG25 and EG47 retained 55-60% of the maximum activity at pH 8.5. Substrate specificity of the purified cellulases against carboxymethylcellulose (CMC), beta-glucan, Avicel, xylan, xyloglucan, laminarin, and p-nitrophenyl-beta-D-cellobioside was studied. EG44 and EG51 were characterized by the highest CMCase activity (59 and 52 U/mg protein). EG28 had the lowest CMCase activity (11 U/mg) amongst the endoglucanases; however, this enzyme displayed the highest activity against beta-glucan (125 U/mg). Only EG51 and CBH I were characterized by high adsorption ability on Avicel cellulose (98-99%). Kinetics of Avicel hydrolysis by the isolated cellulases in the presence of purified beta-glucosidase from Aspergillus japonicus was studied. The hydrolytic efficiency of cellulases (estimated as glucose yield after a 7-day reaction) decreased in the following order: CBH I, EG60, CBH II, EG51, EG47, EG25, EG28, EG44.

MeSH terms

  • Adsorption
  • Cellulase / chemistry
  • Cellulase / isolation & purification*
  • Cellulose 1,4-beta-Cellobiosidase / chemistry
  • Cellulose 1,4-beta-Cellobiosidase / isolation & purification*
  • Chemical Fractionation
  • Chromatography, Gel
  • Chromatography, High Pressure Liquid
  • Chromatography, Ion Exchange
  • Chrysosporium / enzymology*
  • Hydrolysis
  • Isoelectric Focusing
  • Multienzyme Complexes / chemistry
  • Multienzyme Complexes / isolation & purification
  • Mutation / physiology

Substances

  • Multienzyme Complexes
  • Cellulase
  • Cellulose 1,4-beta-Cellobiosidase