Effect on product specificity of cyclodextrin glycosyltransferase by site-directed mutagenesis.
Cyclodextrin glycosyltransferase (CGTase; EC 2.4.1.19) catalyzes the degradation of starch into cyclodextrins through an intramolecular transglycosylation reaction. Tyr-89, Asn-94, and Tyr-100 are located near the putative active center. To analyze their roles in product specificity, Tyr-89, Asn-94, and Tyr-100 of CGTase from alkalophilic Bacillus sp. I-5 were replaced with different amino acids. Among the mutants, the N94S mutant protein produced about two times more alpha-cyclodextrin than the wild-type at all incubation times. The Y89F and Y100F mutant proteins were changed to more beta-specific enzymes. From these results it is suggested that the changing of the residues located at the near active site can change the product specificity of CGTase.
Kim
Y H
YH
Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Yusung-Gu, Taejon, Korea.
Bae
K H
KH
Kim
T J
TJ
Park
K H
KH
Lee
H S
HS
Byun
S M
SM
D000595:Amino Acid Sequence; D001407:Bacillus / Q000201:enzymology*; D001483:Base Sequence; D005964:Glucosyltransferases / Q000737:chemistry*; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D013379:Substrate Specificity
D005964:Glucosyltransferases; C019963:cyclomaltodextrin glucanotransferase