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171.
Fujii K Minagawa H Terada Y Takaha T Kuriki T Shimada J Kaneko H 《Journal of Bioscience and Bioengineering》2007,103(2):167-173
Amylomaltase from Thermus aquaticus catalyzes three types of transglycosylation reaction, as well as a weak hydrolytic reaction of alpha-1,4 glucan. From our previous study [Fujii et al., Appl. Environ. Microbiol., 71, 5823-5827 (2005)], tyrosine 54 (Y54) was identified as an amino acid controlling the reaction specificity of this enzyme. Since Y54 is not located around the active site but in the proposed second glucan binding site that is 14 A away from catalytic residues, the functions of Y54 and the second glucan binding site are of great interest. In this study, we introduced mutations into another tyrosine (Y101) in the second glucan binding site. The obtained mutated enzymes were subjected to all four types of enzyme assay and the effects of mutations on the reaction specificities of these enzymes were comprehensively investigated. These studies indicated that the amino acid substitution at Y54 or Y101 for removing their aromatic side chain increases cyclization activity (intra-molecular transglycosylation reaction) but decreases disproportionation, coupling and hydrolytic activities (inter-molecular reactions). The superimposition of the reported structures of the enzyme with and without substrate analog revealed the occurrence of a conformational change in which a donor binding site becomes open. From lines of evidence, we conclude that the binding of glucan substrate to the second glucan binding site through an interaction with the aromatic side chains of Y54 and Y101 is a trigger for the enzyme to take a completely active conformation for all four types of activity, but prevents the cyclization reaction to occur since the flexibility of the glucan is restricted by such binding. 相似文献
172.
Starch granules were isolated from rice endosperms of twenty mutants, included two kinds of shrunken (shr) mutant, and two cultivars of their normal counterparts. The shr mutants, EM-20 and Kenkei 2088, showed opposite effects on the amylose content, that is, EM-20 had 5% higher than its normal counterpart and Kenkei 2088 had 3% lower than its one. The amylopectin of EM-20 was composed of slightly shorter chains on the average than its normal one. The gelatinization temperature and heat of gelatinization were affected by the mutation of shr, EM-20. The amylose extender (ae) mutant, Kenkei 2064, had a comparable amylose content with its normal counterpart but its amylopectin had higher content of long chain than the normal one. Twelve kinds of dull (du) mutant reduced the amylose content which varied from 2% to 16%. 相似文献
173.
Naoyoshi Inouchi Hideaki Ando Masako Asaoka Kazutoshi Okuno Hidetsugu Fuwa 《Starch - St?rke》2000,52(1):8-12
The effects of environmental temperature during the early development of seeds on the structural characteristics of the endosperm starch were investigated using near‐isogenic lines of rice plants (Taichung 65, waxy), grown under temperature controlled conditions. High performance gel permeation chromatography (HPLC) demonstrated that Pseudomonas isoamylase‐debranched amylopectins of rice plants grown at lower temperature (25°C) contain increased amounts of short chains and decreased amounts of long chains as compared with amylopectins obtained from rice plants grown at higher temperature (30°C). By high performance anion exchange chromatography with pulsed amperometric detection (HPAEC‐PAD) of isoamylase‐debranched amylopectins it was detected that the amount of unit chains with degree of polymerization (DP) 6 and 11‐13, in the amylopectin of rice plants grown at lower temperature (25°C) had significantly increased and the amount of unit chains with DP 8, 22‐24 and 29 had significantly decreased, as compared with the amylopectin of rice plants grown at higher temperature (30°C). It was confirmed that the environmental temperature between 5 and 10 d after pollination strongly influenced the structure characteristics of the endosperm starch of rice plants. 相似文献