Increase in Endoglucanase Productivity and Mycelial Stability of Rhizopus oryzae by Classical Mutagenesis

Aim: To develop a mutant strain with high endoglucanase productivity and optimization of some cultivation parameters.
Place and Duration of Study: Microbiology Research Laboratory, Department of Zoology, Molecular Biology & Genetics, Presidency University, College Street, Kolkata: 700 073, India, between Aug, 2010 and March 2011.
Methodology: The wild strain of Rhizopus oryzae PR7 MTCC 9642 was subjected to classical mutagenesis by suspending 5 hyphal discs (0.5 cm) in 10ml of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) solutions of various concentrations (125-1000µg). The in situ cellulolytic activity of the colonies of the mutant strains on the plates were measured by using alcoholic iodine solution and the highest enzyme producing mutant was selected. The mutant strain was later cultivated in presence of various domestic wastes at various pH, temperature, time. The morphological alteration was also checked by staining with fluorescent dye.
Results: Out of 50 mutants, strain A7 was selected that showed about 33% increase in endoglucanase synthesis utilizing orange bagasse as sole carbon source in a shake flask screen. The strain was found to have the same pH and temperature optima, but could achieve highest level of enzyme production earlier than that by its wild counterpart. Being a dimorphic fungus, the wild type strain of Rhizopus oryzae, showed a transformation to yeast like pelleted form, whereas the mutant strain A 7 showed persistent filamentous structure indicating the achievement of a structural stability in presence of environmental stress.
Conclusion: The present mutant strain could ferment orange bagasse and showed an increased production of endoglucanase with minimized time consumption with greater mycelial stability against various environmental stresses. These achievements will definitely add economy in industrial production of endoglucanse at a nominal cost.