| ?-Glucosidase from Aspergillus niger(A.niger)hydrolyzes ?-1,4 glycosidic bond in starch and is the key enzyme in the production process of isomaltooligosaccharide.Seven genes are predicted to be putative ?-glucosidase genes in the annotated A.niger CBS513.88 genome,however,the gene encoding the predominant ?-glucosidase in the fermentation process has not been reported.In this study,a set of ?-glucosidase gene deletion mutants were constructed using homologous recombination based method.Residue ?-glucosidase activities of the ?-glucosidase gene deletion mutants were measured.Results showed that among the gene deletion mutants,compared with the control strain,only the agdB deletion mutant showed a decrease in ?-glucosidase activity(36%),and no significant decrease were observed in the other gene deletion mutants.Each of agdB and agd A genes was deleted or overexpressed,respectively,in A.niger SH-2-3 which is a glucoamylase and amylase deficient strain.The agd B deletion mutant grew the most slowly in glucose drop-out medium,and the growth rate increased as the concentration of glucose in the medium increases.In the agdB overexpressing strain,the ?-glucosidase activity increased by 10-fold compared with the host strain,while no significant impact on the ?-glucosidase activity was observed in the agdA overexpressing strain.These results further indicated that agdB is the most important functional gene among all the seven predicted ?-glucosidase genes studied in this study.The agdB product is accountable for 36% of the total ?-glucosidase activity in A.niger,and the other six predicted ?-glucosidase genes showed no observable impact on the total ?-glucosidase activity in A.niger SH-2.Taking into account the fact that most(64%)of the total ?-glucosidase activity does not come from the seven predicted ?-glucosidase genes,new genes that encode for proteins with ?-glucosidase activity remain to be further discovered and investigated. |
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