Change in maltose- and soluble starch-hydrolyzing activities of chimeric alpha-glucosidases of Mucor javanicus and Aspergillus oryzae.
The chimeric alpha-glucosidases of Mucor javanicus and Aspergillus oryzae, which has high activity toward not only maltooligosaccharides but also soluble starch and has high activity toward maltooligosaccharides but faint activity toward soluble starch, respectively, were constructed by shuffling the C-terminal regions where low homology is observed between the two enzymes. The chimera genes were expressed in Pichia pastoris to produce and secrete the enzymes that have predicted molecular masses in the culture medium. The two chimeric M. javanicus alpha-glucosidases, of which the N- and C-terminal regions are substituted for those of A. oryzae, respectively, decreased in soluble starch-hydrolyzing activity, however, increased in maltose-hydrolyzing activity by 2.1 and 4.9 times higher than that of the native form of M. javanicus alpha-glucosidase, respectively. The chimeric enzymes changed on the V(max) values for maltose significantly, whereas the K(m) values were similar to that of the native enzyme.
Sugimoto
Manabu
M
Research Institute for Bioresources, Okayama University, 2-20-1 Chuo, Okayama 710-0046, Kurashiki, Japan. manabus@rib.okayama-u.ac.jp
Ohta
Takeshi
T
Kawai
Fusako
F
D000595:Amino Acid Sequence; D001236:Aspergillus oryzae / Q000201:enzymology* / Q000235:genetics; D001665:Binding Sites; D008320:Maltose / Q000378:metabolism; D008969:Molecular Sequence Data; D009089:Mucor / Q000201:enzymology* / Q000235:genetics; D009844:Oligosaccharides / Q000378:metabolism; D010843:Pichia / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D013213:Starch / Q000378:metabolism; D013379:Substrate Specificity; D000520:alpha-Glucosidases / Q000096:biosynthesis / Q000378:metabolism*
D009844:Oligosaccharides; D011993:Recombinant Fusion Proteins; C021705:maltooligosaccharides; D008320:Maltose; D013213:Starch; D000520:alpha-Glucosidases