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Studies On A Thermostable α-Glucosidase From Hyperthermophilic Archaea Thermococcus Siculi HJ21

Posted on:2009-08-19Degree:MasterType:Thesis
Country:ChinaCandidate:L YangFull Text:PDF
GTID:2120360272456690Subject:Microbiology
Abstract/Summary:PDF Full Text Request
Alpha-glucosidase (EC 3.2.1.20), also namedα-transglucosidase and maltase, could catalyze the liberation ofα-D-glucose from the non-reducing end of poly- and oligo-saccharides. It also catalyses transglucosidation reaction to synthesis isomerized oligosaccharides. Alpha-glucosidase was generally involved in the last step of starch degradation. It was used to produce high glucose syrup and iso-maltooligosaccharide withα-amylase in the industrial starch processing. At present, most ofα-glucosidase used in industry was from Aspergillus niger.The culture condition forα-glucosidase production with Thermococcus siculi HJ21 was optimized and the characters of crudeα-glucosidase from T. siculi HJ21 were studied. The optimal culture time for intracellular and extracellularα-glucosidase production was around 6 h and 27 h respectively. The optimal temperature, pH and NaCl concentration forα-glucosidase production were 85℃, 6.5 and 2.5 %, respectively. Addition of soluble starch, yeast extract and peptone to the fermentation media enhanced enzyme production. The optimum temperature for enzyme reaction was 100℃, and the enzyme exhibited extreme thermostability, with a half-life of 2 h at 90℃. Theα-glucosidase exhibited maximum activity at pH 7.0, and was stable at pH 5.0~8.0. The addition of Ca2+ was not benefit toα-glucosidase thermostability. The enzyme activity was significantly inhibited by Cu2+, Al3+and Ni2+. Hg2+completely inhibits the T. siculi HJ21α-glucosidase activity. The enzyme activity was activatied by EDTA, Fe3+and K+.Theα-glucosidase gene was cloned successfully from T. siculi HJ21 with PCR method. The nucleotide sequence analysis revealed that theα-glucosidase has a 729 bp open reading frame, which encodes a protein of 242 amino acid residue with the molecular weight of about 27.2 kDa. According to the GeneBank database, the homology ofα-glucosidase gene from T. siculi HJ21 shares 81% with T. hydrothermalis, and the similarity of amino acid sequences was 90%.
Keywords/Search Tags:Thermophilic Archaeon, Thermococcus siculi HJ21, Thermophilic enzyme, α-Glucosidase
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