ISSN: 0973-7510
E-ISSN: 2581-690X
A starch-binding domain encoding region of glucoamylase (GASBD) was amplified by PCR from Aspergillus niger 2316 and sequenced (GenBank JQ814697). It consists of 324bp nucleotides and encodes a peptide of 108 amino acids. To investigate the affinities of artificial multiple-repeat GASBDs for insoluble starch in vitro, the pET28a plasmids harbouring, respectively, single GASBD and multiple-repeat GASBDs from two to four were constructed and expressed in Escherichia coli BL21(DE3). The recombinant GASBD(s) proteins were purified to homogeneity by Ni2+–NTA affinity chromatography and the affinities of the purified recombinant proteins for insoluble starches were determined. GASBD2, GASBD3, and GASBD4 had, respectively, approximately 5, 8 and 10-fold higher affinities for starch than single GASBD, indicating that the multiple-repeat GASBDs act in a synergistic way when binding to starches.
Glucoamylase, Starch-binding domain, Protein expression, Starch affinity, Synergistic effect
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