Abstract
Two strains of Trichoderma reesei, wild type QM6a and mutant Rut-C30, were grown in meida containing an inducer, insoluble crystalline cellulose (Avicel PH101), as carbon source for 11 days. The cell growth, expressed as myceliar protein content, of Rut-C30 was 4–5 times higher than QM6a. The lack of ultrastructural disorganization, and absence of intracellular enzyme release into the growth medium, indicated that none of these two strains had undergone any significant autolysis during the entire growth phase. Cellulase activities, mainly endoglucanase, cellobiase and filter paper degrading activity (disc) were enhanced in Rut-C30 cells. A major change was observed in the endoglucanase activity which was 30 times higher in Rut-C30 than QM6a, whereas, both β-glucosidase and disc activities were 3 times enhanced in Rut-C30 compared to QM6a. In addition to synthesis, cellulase secretion was also enhanced in Rut-C30. Both the organisms contained same amounts of intracellular marker enzyme activities (e.g., inosine diphosphatase, thiamine pyrophosphatase, alkaline phosphatase). Finally, the enahncement of secretory activity of Rut-C30 was correlated with the proliferation of rough endoplasmic reticulum (RER) and increased phospholipid content. It appears that Rut-C30 is not only a hypercellulolytic but also a hypersecretor mutant.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Bauer H, Sigarlakie E (1975) Localization of alkaline phosphatase in Saccharomyces cerevisiae by means of ultrathin frozen sections. J Ultrastruct Res 50:208–215
Brown RD Jr, Jurasek L (Ed) (1979) Hydrolysis of cellulose: Mechanism of enzymatic and acid catalysis. Amer Chem Soc, Adv Chem Ser 181
Chrispeels MJ (1976) Biosynthesis, intracellular transport, and secretion of extracellular macromolecules. Ann Rev Plant Physiol 27:19–38
Dahlquest A (1964) Methods for assay intestinal disaccharidases. Anal Biochem 7:18–25
Dillon LS (1981) Ultrastructure, macro-molecules and evolution, p 201–316, Plenum Pres, New York
Dittmer JC, Dawson RMC (1961) The isolation of a new lipid triphosphoinositide and monophosphoinositide from Ox brain. Biochem J 81:535–540
Ernster L, Zetterstrom R, Lindberg O (1950) A method for the determination of tracer phosphate in biological material. Acta Chem Scand 4:942–947
Eveleigh DE, Montenecourt BS (1979) Increasing yields of extracellular enzymes. Adv in Appl Microbiol 25:57–74
Gascoigne JA, Gascoigne MM (1960) Biological degradation of cellulose. Butterworths and Co. London
Ghosh BK (1977) Techniques to study the ultrastructure of microorganisms. In: Laskin AI, Lechevalier HA (eds) Handbook of Microbiology, Vol 1, Bacteria. CRC Press, Ind Ohio, pp 31–38
Ghosh A, Ghosh BK (1972) Changes in the membrane bound alkaline phosphatase of glucose and lactate grown vegetative cells of Bacillus subtilis SB15. Biochem Biophys Res Commun 49:906–915
Ghosh A, Al-Rabiai S, Ghosh BK, Trimino-Vazquez H, Eveleigh DE, Montenecourt BS (1982) Increased endoplasmic reticulum content of a mutant of Trichoderma reesei (Rut-C30) in relation to cellulase synthesis. Enzyme Microb Technol 4:110–113
Ghosh A, Glenn MG, Ghosh BK (1983) Polymorphism of endoplasmic reticulum organization in a hypercellulolytic mutant of Trichoderma veesei Rut-C30. Abst. Am Soc Cell Biol P-305a
Gooday GW, Trinci APJ (1980) Wall structure and biosynthesis in fungi. In: Symp Soc Gen Microbiol 30:207–251
Grove SN, Bracker CE (1970) Protoplasmic organization of hyphal tips among fungi: vesicles and spitzenkorper. J Bacteriol 104:989–1009
Hand AR, Oliver C (1981) The Golgi Apparatus: Protein transport and packaging in secretory cells. In: Hand AR, Oliver C (eds) Methods in Cell Biology. Academic Press, New York 23:137–153
Handley D, Ghosh BK (1980) Subcellular distribution of marker enzymes in cells of a minute fungus, Fusidium sp 100-3. J Bacteriol 141:946–955
Li LH, Flora RM, King KW (1965) Individual roles of cellulase components derived from Trichoderma viride. Arch Biochem Biophys 111:439–447
Lloyd JB, Whelan WJ (1969) An improved method for enzymic determination of glucose in the presence of maltose. Anal Biochem 30:467–470
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurements with the Folin phenol reagent. J Biol Chem 193:265–275
Mandels M, Weber J (1969) The production of cellulase. In: Cellulases and their application. Adv Chem Ser 95:391–414
Mandels M, Hontz L, Nystrom J (1974) Enzymatic hydrolysis of waste cellulose. Biotechnol Bioeng 16:1471–1493
Mandels M, Andreotti R, Roche C (1976) Measurement of saccharifying cellulase. In: Gaden EL, Mandels MH, Reese ET, Spano LA (eds) Enzymatic conversion of cellulosic material: technology and applications. John Wiley and Sons, New York, p 21–33
Miller GL (1969) Use of dinitrosalicyclic acid reagent for determination of reducing sugar. Anal Chem 31:426–428
Montenecourt BS, Eveleigh DE (1979) Selective screening methods for the isolation of high yielding cellulase mutants of T. reesei. In: Brown RD Jr, Jurasek L (eds) Hydrolysis of cellulose: Mechanisms of enzymatic and acid catalysis. Adv Chem Ser 181:289–301
Morre DJ, Ovtract L (1977) Dynamics of Golgi apparatus. Membrane differentiation and membrane flow. Int Rev Cytol (suppl) 5:61–188
Nisizawa T, Suzuki H, Nisizawa K (1971) “De nova” synthesis of cellulase induced by sophorose in Trichoderma viride cells. J Biochem 70:387–393
Novick P, Ferro S, Schekman R (1981) Order of events in yeast secretory pathyway. Cell 25:461–469
Novikoff AB (1976) The endoplasmic reticulum. A cytochemist's view (A review). Proc Natl Acad Sci 73:2781–2787
Novikoff AB, Essner E, Goldfischer S, Heus M (1962) Nucleoside-phosphatase activities of cytomembranes. Symp Intern Soc Cell Biol 1:149–192
Reynold ES (1963) The use of lead citrate at high pH as an electron opaque stain in electron microscopy. J Cell Biol 17:208–212
Schall BF, Marathe GV, Ghosh BK (1981) Stereological analysis of plasmolysis in logarithmic phase Bacillus licheniformis. J Bacteriol 146:391–397
Selby K, Maitland CC (1967) The cellulase of Trichoderma viride — separation of the components involved in the solubilization of cotton. Biochem J 104:716–724
Shoemaker SP, Raymond JC, Brunos R (1981) Cellulases: Diversity amongst improved Trichoderma strains. In: Hollaender A (ed) Trends in the Biology of Fermentations for Fuels and Chemicals. Plenum Press, New York 18:89–109
Vogel HJ (1956) A convenient growth medium for Neurospora (medium N). Microb Genet Bull 13:42–43
Weibel ER (1969) Stereological principles for morphometry in electron microscopic cytology. Int Rev Cytol 26:235–502
White AR, Brown RM Jr (1981) Enzymatic hydrolysis of cellulose: Visual characterization of the process. Proc Natl Acad Sci 78:1047–1051
Wood TM (1969) The cellulase of Fusarium solani — resolution of the enzyme complex. Biochem J 115:457–464
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Ghosh, A., Ghosh, B.K., Trimino-Vazquez, H. et al. Cellulase secretion from a hyper-cellulolytic mutant of Trichoderma reesei Rut-C30. Arch. Microbiol. 140, 126–133 (1984). https://doi.org/10.1007/BF00454914
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00454914