Overproduction of glucoamylase by a deregulated mutant of a thermophilic mould Thermomucor indicae-seudaticae.

Thermomucor indicae-seudaticae, a glucoamylase-producing thermophilic mould, was mutagenised using nitrous acid and gamma ((60)Co) irradiation in a sequential manner to isolate deregulated mutants for enhanced production of glucoamylase. The mutants were isolated on Emerson YpSs agar containing a non-metabolisable glucose analogue 2-deoxy-D-glucose (2-DG) for selection. The preliminary screening for glucoamylase production using starch-iodine plate assay followed by quantitative confirmation in submerged fermentation permitted the isolation of several variants showing varying levels of derepression and glucoamylase secretion. The mutant strain T. indicae-seudaticae CR19 was able to grow in the presence of 0.5 g l(-1) 2-DG and produced 1.8-fold higher glucoamylase. As with the parent strain, glucoamylase production by T. indicae-seudaticae CR19 in 250-ml Erlenmeyer flasks attained a peak in 48 h of fermentation, showing higher glucoamylase productivity (0.67 U ml(-1) h(-1)) than the former (0.375 U ml(-1) h(-1)). A large-scale cultivation in 5-l laboratory bioreactor confirmed similar fermentation profiles, though the glucoamylase production peak was attained within 36 h attributable to the better control of process parameters. Although the mutant grew slightly slow in the presence of 2-DG and exhibited less sporulation, it showed faster growth on normal Emerson medium with a higher specific growth rate (0.138 h(-1)) compared to the parent strain (0.123 h(-1)). The glucoamylase produced by both strains was optimally active at 60 degrees C and pH 7.0 and displayed broad substrate specificity by cleaving alpha-1,4- and alpha-1,6-glycosidic linkages in starch, amylopectin, amylose and pullulan. Improved productivity and higher specific growth rate make T. indicae-seudaticae CR19 a useful strain for glucoamylase production.