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PubMed2005

Sorption and immobilization of cellulase on silicate clay minerals.

The interaction of organic molecules with mineral surfaces is a subject of interest in a variety of disciplines. Enzymes are able to be sorbed and immobilized by clay minerals and humic colloids in soil environment. The present study was done to elucidate some aspects of sorption and immobilization of cellulase on soil components by analysis of the sorption, and immobilization of cellulase on Avicel, a soil sample, illite, kaolinite, montmorillonite, and palygorskite. Palygorskite displayed the highest sorption capacity. Sorbents coated with hydroxyaluminum displayed significantly higher capacity than uncoated sorbents. The positive effects of Al(OH)(x) coating on sorption capacities of the different sorbents were not equal. The effect decreased in the order soil > palygorskite > kaolinite > Avicel > montmorillonite > illite. The amount of sorbed cellulase desorbed from external surfaces of soil was quite low (about 16%), especially in coated samples (about 6%). X-ray diffraction analysis of K-montmorillonite and Ca-montmorillonite showed that Al(OH)(x) was intercalated between the montmorillonite layers. Immobilization of cellulase on the sorbents did not result in expansion of their crystal structures. Therefore, it may be concluded that the amount of cellulase immobilized on internal surfaces of the sorbents was negligible.

Adsorption↗
PubMed2000

Purification and characterization of cellulases produced by two Bacillus strains.

Cellulases produced by two Bacillus strains, CH43 and HR68, isolated from hot springs in Zimbabwe, were purified to homogeneity from culture supernatants. Both enzymes had molecular mass of 40 kDa and isoelectric point of 5.4. The enzymes also resembled each other in N-terminal amino acid sequence which was Ala-Gly-Thr-Lys-Thr-Pro-Val-Ala-Lys-Asn-Gly-Gln, showing 100% homology with that of endoglucanases from Bacillus subtilis belonging to glycoside hydrolase family five. The cellulases were optimally active in the pH range of 5-6.5. The optimum temperature was 65 and 70 degrees C for the endoglucanase of CH43 and HR68, respectively. The CH43 enzyme was stable at 50 degrees C in a pH range of 6-10, and HR68 at pH 6-8. Both the enzymes retained complete activity for at least 24 h at 50 degrees C. The enzymes showed highest activity with beta-glucan as substrate followed by carboxymethylcellulose. Significant activity was also observed with crystalline forms of cellulose such as filter paper and Avicel, particularly for HR68 cellulase. For carboxymethycellulose, the CH43 and HR68 cellulases had a Km of 1.5 and 1.7 mg ml(-1), respectively, and Vmax of 0.93 and 1.70 mmol glucose min(-1) mg protein(-1) respectively. The activity of the enzymes was not influenced by most metal ions at 1 mM concentration, but was increased by about 38% by Co2+. The inhibition by Hg2+ and Mn2+ was higher for CH43 than for HR68 enzyme. Ag+ inhibited the CH43 activity but stimulated the HR68 activity. The CH43 cellulase was inhibited by N-bromosuccinimide and iodoacetamide while HR68 was unaffected.

Bacillus↗
PubMed2001

Biostoning of denims by Penicillium occitanis (Pol6) cellulases.

The Pol6 mutant of Penicillium occitanis, secreting a large quantity of cellulases, was cultivated in fermentor using a local paper pulp as an inducer substrate. A high titer of extracellular cellulase activity was reached after a fed batch process: 23 IU x ml(-1) filter paper activity, 21 IU x ml(-1) CMCases activity (endoglucanase units) and 25 mg x ml(-1) of proteins. Various tests were done to compare the action of the P. occitanis cellulases with those commercially available and with the traditional stonewashing process. This cellulase preparation was successfully applied in a biostoning process at an industrial scale. The abrasive effect of the P. occitanis cellulases was very uniform and with an efficiency comparable to that obtained by the commercial ones.

Cellulase↗
PubMed1997

Kinetic model for batch cellulase production by Trichoderma reesei RUT c30.

A kinetic model for batch cellulase enzyme production by T. reesei from cellulose substrate is constructed from literature concepts and laboratory data. The key concepts included were four: (i) existence of primary and secondary mycelia; (ii) cellulase production by secondary mycelia only; (iii) the adsorption of cellulase (catalyst) on the particulate cellulose (substrate), and (iv) the decline of cellulose reactivity with extent of conversion. The laboratory batch data were biomass (particulate), substrate (particulate cellulose), and product (cellulase enzyme and reducing sugar) concentration vs. time. The kinetic parameters were evaluated simultaneously through a nonlinear fitting routine, and the resultant model is shown to fit the data well. The model's success validates the presumed need to include all four concepts in reactor analysis for cellulase production.

Adsorption↗
PubMed1979

Production of cellulase (Cx) by different species of Erwinia.

The tested isolates of Erwinia chrysanthemi (corn pathotype) and E. carotovora constitutively produce high levels of cellulase(s) (Cx) in presence or absence of carboxymethyl-cellulose (CMC) as substrate in the medium. The tested isolates of E. atroseptica produced high levels of cellulase when grown in presence of both carboxymethyl-cellulose (CMC) and sucrose, low levels in presence of carboxymethyl-cellulose alone, and traces of cellulase(s) in presence of sucrose alone. The activity of cellulase(s), present in culture supernatants of E. chrysanthemi (corn pathotype) and E. carotovora, occurred in a broad range of pH value (2.2--9) with an optimum pH ranging from pH 4 to 7. However, the activity of cellulase(s), present in culture supernatant of E. atroseptica, occurred in a narrow range of pH value (3--7) with an optimum of about pH 5.

Carboxymethylcellulose Sodium↗
PubMed1979

The characterization of cellulase from Paecilomyces fusisporus Saksena.

The fractionation of cellulase from Paecilomyces fusisporus Saksena on DEAE-Sephadex (A-5O) resulted in separation of beta-glucosidase, cellulase (on cellulose powder), and CM cellulase activity. Cellulase activity was associated with some CM cellulase activity, whereas the latter was independent of the former.

Carboxymethylcellulose Sodium↗
PubMed2002

4-Methyl-7-thioumbelliferyl-beta-D-cellobioside: a fluorescent, nonhydrolyzable substrate analogue for cellulases.

The kinetics of cellulose binding and hydrolysis by cellulases is not well understood except at steady-state conditions. For use in studies of cellulase pre-steady-state and steady-state kinetics, we have prepared 4-methyl-7-thioumbelliferyl-beta-D-cellobioside (MUS-CB), a ground-state nonhydrolyzable analogue of the fluorescent cellulase substrate 4-methylumbelliferyl-beta-D-cellobioside (MU-CB). MUS-CB is not hydrolyzed by the catalytic domain of cellulase E1 from Acidothermus cellulolyticus under conditions where this enzyme rapidly degrades MU-CB. Thermodynamic parameters describing the steady-state binding of MUS-CB to Thermobifida fusca cellulase Cel6A are similar to those for MU-CB, indicating that MUS-CB can be used in place of MU-CB to study binding events in the Cel6A active-site cleft. In the pre-steady-state, MUS-CB binds to Cel6A by a simple, one-step bimolecular association reaction. It is anticipated that similar thio-containing 4-methylumbelliferyl compounds will have applications in studies of other enzyme systems.

Binding Sites↗
PubMed2003

Enzymic release of reducing sugars from oat hulls by cellulase, as influenced by Aspergillus ferulic acid esterase and trichoderma xylanase.

Hydroxycinnamic acids, mainly ferulic and p-coumaric acids, are believed to be inhibitory to ruminal biodegradability of complex cell wall materials such as oat hulls. Previous studies have shown that a novel enzyme, Aspergillus ferulic acid esterase, and Trichoderma xylanase act synergistically to break the ester linkage between ferulic acid and the attached sugar of feruloyl polysaccharides, releasing ferulic acid from oat hulls. In this paper, we examined the enzymic release of reducing sugars from oat hulls by the actions of individual enzymes (Aspergillus ferulic acid esterase at 13 mU, 6.4 U, and 4678.4 U/assay; cellulase at 20 levels, ranging from 7.8 mU to 2772.7 U/assay; Trichoderma xylanase at 20 levels, ranging from 7.8 mU to 4096 U/assay) and by the combined action of cellulase at six levels (62.5 mU, 2 U, 16 U, 128 U, 1024 U, and 2772.7 U/assay), Aspergillus ferulic acid esterase at 13 mU/assay, and Trichoderma xylanase at two levels (1 U and 256 U/assay). The amount of total acid-extractable reducing sugars in the oat hulls used in this study was 793.8 +/- 8.0 microg/mg. The results show that after a 24-h incubation with Aspergillus ferulic acid esterase alone, no reducing sugars were observed to be released from oat hulls. With cellulase as the sole enzyme, as the concentration increased from 7.8 mU to 2772.7 U/assay, the release of reducing sugars increased (P < 0.01) from 0 to 39% of the total present, with the highest release at 512 U/assay. With Trichoderma xylanase alone, as the concentration increased from 7.8 mU to 4096 U/assay, the release of reducing sugars increased (P < 0.01) from 4.9 to 33%, with the highest release at 2048 U/assay. When incubated together with Trichoderma xylanase (1 U or 256 U/assay) and Aspergillus ferulic acid esterase (13 mU/assay), cellulase at all six levels (62.5 mU, 2 U, 16 U, 128 U, 1024 U and 2772.7 U/assay) significantly increased the release of reducing sugars (P < 0.01) from 8 to 69%. These results indicate that the synergistic interaction between Aspergillus ferulic acid esterase and Trichoderma xylanase on the release of ferulic acid from feruloyl polysaccharides of oat hulls makes the remainder of the polysaccharides open for further hydrolytic attack and facilitates the accessibility of the main chain of polysaccharides to cellulase. This action extends the cell wall hydrolysis, thus releasing a higher yield of reducing sugars. Such enzymic pretreatment of oat hulls may provide a unique advantage to rumen microorganisms for the biodegradation of the complex cell walls of byproduct feeds such as oat hulls.

Aspergillus↗
PubMed2004

Kinetic and structural optimization to catalysis at low temperatures in a psychrophilic cellulase from the Antarctic bacterium Pseudoalteromonas haloplanktis.

The cold-adapted cellulase CelG has been purified from the culture supernatant of the Antarctic bacterium Pseudoalteromonas haloplanktis and the gene coding for this enzyme has been cloned, sequenced and expressed in Escherichia coli. This cellulase is composed of three structurally and functionally distinct regions: an N-terminal catalytic domain belonging to glycosidase family 5 and a C-terminal cellulose-binding domain belonging to carbohydrate-binding module family 5. The linker of 107 residues connecting both domains is one of the longest found in cellulases, and optimizes substrate accessibility to the catalytic domain by drastically increasing the surface of cellulose available to a bound enzyme molecule. The psychrophilic enzyme is closely related to the cellulase Cel5 from Erwinia chrysanthemi. Both kcat and kcat/K(m) values at 4 degrees C for the psychrophilic cellulase are similar to the values for Cel5 at 30-35 degrees C, suggesting temperature adaptation of the kinetic parameters. The thermodynamic parameters of activation of CelG suggest a heat-labile, relatively disordered active site with low substrate affinity, in agreement with the experimental data. The structure of CelG has been constructed by homology modelling with a molecule of cellotetraose docked into the active site. No structural alteration related to cold-activity can be found in the catalytic cleft, whereas several structural factors in the overall structure can explain the weak thermal stability, suggesting that the loss of stability provides the required active-site mobility at low temperatures.

Amino Acid Sequence↗
PubMed2004

The galactokinase of Hypocrea jecorina is essential for cellulase induction by lactose but dispensable for growth on d-galactose.

Lactose is the only soluble carbon source which can be used economically for the production of cellulases or heterologous proteins under cellulase expression signals by Hypocrea jecorina (=Trichoderma reesei). Towards an understanding of lactose metabolism and its role in cellulase formation, we have cloned and characterized the gal1 (galactokinase) gene of H. jecorina, which catalyses the first step in d-galactose catabolism. It exhibits a calculated Mr of 57 kDa, and shows moderate identity (about 40%) to its putative homologues of Saccharomyces cerevisiae and Kluyveromyces lactis. Gal1 is a member of the GHMP family, shows conservation of a Gly/Ser rich region involved in ATP binding and of amino acids (Arg 51, Glu 57, Asp 60, Asp 214, Tyr 270) responsible for galactose binding. A single transcript was formed constitutively during the rapid growth phase on all carbon sources investigated and accumulated to about twice this level during growth on d-galactose, l-arabinose and their corresponding polyols. Deletion of gal1 reduces growth on d-galactose but does only slightly affect growth on lactose. This is the result of the operation of a second pathway for d-galactose catabolism, which involves galactitol as an intermediate, and whose transient concentration is strongly enhanced in the delta-gal1 strain. In this pathway, galactitol is catabolised by the lad1-encoded l-arabinitol-4-dehydrogenase, because a gal1/lad1 double delta-mutant failed to grow on d-galactose. In the delta-gal1 strain, induction of the Leloir pathway gene gal7 (encoding galactose-1-phosphate uridylyltransferase) by d-galactose, but not by l-arabinose, is impaired. Induction of cellulase gene expression by lactose is also impaired in a gal1 deleted strain, whereas their induction by sophorose (the putative cellulose-derived inducer) was shown to be normal, thus demonstrating that galactokinase is a key enzyme for cellulase induction during growth on lactose, and that induction by lactose and sophorose involves different mechanisms.

Amino Acid Sequence↗
PubMed2003

Purification and cDNA cloning of a cellulase from abalone Haliotis discus hannai.

A cellulase [endo-beta-1,4-D-glucanase (EC 3.2.1.4)] was isolated from the hepatopancreas of abalone Haliotis discus hannai by successive chromatographies on TOYOPEARL CM-650M, hydroxyapatite and Sephacryl S-200 HR. The molecular mass of the cellulase was estimated to be 66 000 Da by SDS/PAGE, thus the enzyme was named HdEG66. The hydrolytic activity of HdEG66 toward carboxymethylcellulose showed optimal temperature and pH at 38 degrees C and 6.3, respectively. cDNAs encoding HdEG66 were amplified by the polymerase chain reaction from an abalone hepatopancreas cDNA library with primers synthesized on the basis of partial amino-acid sequences of HdEG66. By overlapping the nucleotide sequences of the cDNAs, a sequence of 1898 bp in total was determined. The coding region of 1785 bp located at nucleotide position 56-1840 gave an amino-acid sequence of 594 residues including the initiation methionine. The N-terminal region of 14 residues in the deduced sequence was regarded as the signal peptide as it was absent in HdEG66 protein and showed high similarity to the consensus sequence for signal peptides of eukaryote secretory proteins. Thus, matured HdEG66 was thought to consist of 579 residues. The C-terminal region of 453 residues in HdEG66, i.e. approximately the C-terminal three quarters of the protein, showed 42-44% identity to the catalytic domains of glycoside hydrolase family 9 (GHF9)-cellulases from arthropods and Thermomonospora fusca. While the N-terminal first quarter of HdEG66 showed 27% identity to the carbohydrate-binding module (CBM) of a Cellulomonas fimi cellulase, CenA. Thus, the HdEG66 was regarded as the GHF9-cellulase possessing a family II CBM in the N-terminal region. By genomic PCR using specific primers to the 3'-terminal coding sequences of HdEG66-cDNA, a DNA of 2186 bp including three introns was amplified. This strongly suggests that the origin of HdEG66 is not from symbiotic bacteria but abalone itself.

Amino Acid Sequence↗
PubMed2001

ACEII, a novel transcriptional activator involved in regulation of cellulase and xylanase genes of Trichoderma reesei.

A novel yeast-based method to isolate transcriptional activators was applied to clone regulators binding to the cellulase promoter cbh1 of the filamentous fungus Trichoderma reesei (Hypocrea jecorina). This led to the isolation of the cellulase activator ace2 encoding for a protein belonging to the class of zinc binuclear cluster proteins found exclusively in fungi. The DNA-binding domain of ACEII was expressed as a glutathione S-transferase fusion protein in Escherichia coli, and ACEII was shown to bind in vitro to the 5'-GGCTAATAA site present in the cbh1 promoter. This site also contains the proposed binding sequence of the xylanase activator XlnR of Aspergillus niger. Mutation of the GGC triplet abolished ACEII binding. The function of ACEII was studied by analyzing the effects of ace2 deletion in the hypercellulolytic T. reesei strain ALKO2221. Deletion of the ace2 gene led to lowered induction kinetics of mRNAs encoding the major cellulases cellobiohydrolases I and II and endoglucanases I and II and to 30-70% reduced cellulase activity when the fungus was grown on medium containing Solka floc cellulose. The expression level of the gene encoding xylanase was also affected. ace2 deletion led to lowered xyn2 expression in cellulose-induced cultivation. Cellulase induction by sophorose was not affected by ace2 deletion.

Amino Acid Sequence↗
PubMed2002

Cleavage of the xylosyl serine linkage between a core peptide and a glycosaminoglycan chain by cellulases.

We previously found that endo-beta-xylosidase from Patinopecten is an endo-type glycosidase that cleaves the xylosyl serine linkage between a glycosaminoglycan chain and its core protein (Takagaki, K., Kon, A., Kawasaki, H., Nakamura, T., Tamura, S., and Endo, M. (1990) J. Biol. Chem. 265, 854-860). Screening for endo-beta-xylosidase activity in several cellulases detected this activity in the enzymes from Aspergillus niger, Penicillium funiculosum, Trichoderma reesei, Trichoderma viride, and Irpex lacteus. The cellulase derived from A. niger was purified, and its molecular weight was determined to be 26,000 by SDS-PAGE. Examination of the specificity of the cellulase revealed that 1) the enzyme acts on the linkage region (xylosyl serine) between a core peptide and a glycosaminoglycan chain; 2) enzymatic activity is greater with shorter glycosaminoglycan chains; 3) the enzyme readily hydrolyzes the linkage in glycosaminoglycan peptides, but intact proteoglycan is cleaved only slowly; and 4) the activity is unaffected by the glycosaminoglycan component (chondroitin sulfate, dermatan sulfate, and heparan sulfate). Judging from these enzymatic characteristics, this cellulase is different from the endo-beta-xylosidase of Patinopecten. We believe that this cellulase will become a useful tool in the further development of glycotechnology, because, like the endo-beta-xylosidase of Patinopecten, it enables the release of intact glycosaminoglycans from glycosaminoglycan peptides.

Carbohydrate Sequence↗
PubMed1976

Purification and properties of an endo-cellulase of avicelase type from Irpex lacteus (Polyporus tulipiferae).

A culture filtrate of Irpex lacteus (Polyporus tulipiferae) was fractionated initially by salting out with ammonium sulfate, and a cellulase [EC 3.2.1.4.] fraction with high Avicel-hydrolyzing activity (formerly called Avicelase) was extensively purified by a series of column chromatography procedures. This purified endo-cellulase showed a less random hydrolytic mechanism, and was obtained in a yield of 0.04% with respect to the starting material. Its specific activity was enhanced approximately 30 times over that of the starting material. The cellulase component showed a single peak on both ultracentrifugal and acrylamide disc electrophoretic analyses. Its molecular weight was estimated to be 56,000. It contained 12.2% carbohydrate; the major sugar constituents were glucose and mannose. Regarding the amino acid composition, the contents of aspartic acid and glycine were highest, followed by those of glutamic acid, serine, and theonine. The cellulase component was not markedly inhibited by most metal ions tested excepted for Hg2+. This purified endo-cellulase attacked a series of cellooligosaccharides, beta-cellobioside, CM-cellulose, and insoluble, cellulosic substrates. In the digests from insoluble substrates, glucose, cellobiose, cellotriose, and cellotetraose were detectable, but the amount of cellobiose was the largest by far. In constrast, cellobiose and glucose were produced in almost equal amounts from beta-cellobioside.

Amino Acids↗
PubMed2006

Production of cellulase by Aspergillus niger biofilms developed on polyester cloth.

AIMS: To compare cellulase production by Aspergillus niger ATCC 10864 biofilms on polyester cloth and freely suspended cultures in shaken flasks and microbioreactors of bubble column type. METHODS AND RESULTS: Both shaken flasks and oxygenated microbioreactors containing 40 ml of production medium were used to compare cellulase secretion by free mycelium and biofilm cultures. Free mycelium cultures grew better in flasks than in microbioreactors producing compact and fluffy pellets, respectively, while the opposite was found for biofilm cultures without any visible change in biofilm morphology. Cellulase activities and volumetric productivities attained by biofilms in flask cultures were 70% higher than that produced by free mycelium cultures and threefold higher when biofilms were grown in microbioreactors. CONCLUSIONS: Fungal biofilms developed on polyester cloth in both flasks and microbioreactors produce higher cellulase yields and volumetric productivities than free mycelium cultures at lower biomass levels. SIGNIFICANCE AND IMPACT OF THE STUDY: The results of the present study are of commercial and biological interest. All productivity parameters revealed that fungal biofilms may be used for the production of cellulase and other proteins in various types of bioreactors. Moreover, they may be used as model systems to study differential gene expression related to cell adhesion.

Aspergillus niger↗
PubMed1996

Characterization of a Neocallimastix patriciarum cellulase cDNA (celA) homologous to Trichoderma reesei cellobiohydrolase II.

The nucleotide sequence of a cellulase cDNA (celA) from the rumen fungus Neocallimastix patriciarum and the primary structure of the protein which it encodes were characterized. The celA cDNA was 1.95 kb long and had an open reading frame of 1,284 bp, which encoded a polypeptide having 428 amino acid residues. A sequence alignment showed that cellulase A (CELA) exhibited substantial homology with family B cellulases (family 6 glycosyl hydrolases), particularly cellobiohydrolase II from the aerobic fungus Trichoderma reesei. In contrast to previously characterized N. patriciarum glycosyl hydrolases, CELA did not exhibit homology with any other rumen microbial cellulases described previously. Primary structure and function studies in which deletion analysis and a sequence comparison with other well-characterized cellulases were used revealed that CELA consisted of a cellulose-binding domain at the N terminus and a catalytic domain at the C terminus. These two domains were separated by an extremely Asn-rich linker. Deletion of the cellulose-binding domain resulted in a marked decrease in the cellulose-binding ability and activity toward crystalline cellulose. When CELA was expressed in Escherichia coli, it was located predominantly in the periplasmic space, indicating that the signal sequence of CELA was functional in E.coli. Enzymatic studies showed that CELA had an optimal pH of 5.0 and an optimal temperature of 40 degrees C. The specific activity of immunoaffinity-purified CELA against Avicel was 9.7 U/mg of protein, and CELA appeared to be a relatively active cellobiohydrolase compared with the specific activities reported for other cellobiohydrolases, such as T. reesei cellobiohydrolases I and II.

Amino Acid Sequence↗
PubMed2006

Symbiotic "Archaezoa" of the primitive termite Mastotermes darwiniensis still play a role in cellulase production.

The relictual Mastotermes darwiniensis is one of the world's most destructive termites. Like all phylogenetically basal termites, it possesses protozoa in its hindgut, which are believed to help it digest wood. L. Li, J. Frohlich, P. Pfeiffer, and H. Konig (Eukaryot. Cell 2:1091-1098, 2003) recently cloned the genes encoding cellulases from the protozoa of M. darwiniensis; however, they claimed that these genes are essentially inactive, not contributing significantly to cellulose digestion. Instead, they suggested that the protozoa sequester enzymes produced by the termite in its salivary glands and use these to degrade cellulose in the hindgut. We tested this idea by performing gel filtration of enzymes in extracts of the hindgut, as well as in a combination of the salivary glands, foregut, and midgut. Three major cellulases were found in the hindgut, each of which had a larger molecular size than termite-derived salivary gland enzymes. N-terminal amino acid sequencing of one of the hindgut-derived enzymes showed that it was identical to the putative amino acid sequence of one mRNA sequence isolated by Li et al. (Eukaryot. Cell 2:1091-1098, 2003). The overall activity of the hindgut cellulases was found to be of approximately equal magnitude to the termite-derived cellulases detected in the mixture of salivary gland, foregut, and midguts. Based on these results, we conclude that, contrary to Li et al. (Eukaryot. Cell 2:1091-1098, 2003), the hindgut protozoan fauna of M. darwiniensis actively produce cellulases, which play an important role in cellulose digestion of the host termite.

Amino Acid Sequence↗
PubMed1994

Multicomplex cellulase-xylanase system of Clostridium papyrosolvens C7.

The cellulase system of Clostridium papyrosolvens C7 was fractionated by means of ion-exchange chromatography into at least seven high-molecular-weight multiprotein complexes, each with different enzymatic and structural properties. The molecular weights of the complexes, as determined by gel filtration chromatography, ranged from 500,000 to 660,000, and the isoelectric points ranged from 4.40 to 4.85. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of the complexes showed that each complex had a distinct polypeptide composition. Avicelase, carboxymethyl cellulase, and xylanase activity profiles differed from protein complex to protein complex. Three of the complexes hydrolyzed crystalline cellulose (Avicel). Activity zymograms of gels (following electrophoresis under mildly denaturing conditions) revealed different carboxymethyl cellulase-active proteins in all complexes but xylanase-active proteins in only two of the complexes. The xylanase specific activity of these two complexes was more than eightfold higher than that of the unfractionated cellulase preparation. A 125,000-M(r) glycoprotein with no apparent enzyme activity was the only polypeptide present in all seven complexes. Experiments involving recombination of samples eluted from the ion-exchange chromatography column indicated that synergistic interactions occurred in the hydrolysis of crystalline cellulose by the cellulase system. We propose that the C. papyrosolvens enzyme system responsible for the hydrolysis of crystalline cellulose and xylan is a multicomplex system comprising at least seven diverse protein complexes.

Cellulase↗