Crystal structure of a compact ¦Á-amylase from Geobacillus thermoleovorans

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• 作者：Sook-Chen Mok¹ ; Aik-Hong Teh¹ ; ^{aikhong@ccbusm.edu.my} ; Jennifer A. Saito ; Nazalan Najimudin² ; Maqsudul Alam³ ; ⁴ ; ^{alam@hawaii.edu}
• 关键词：GTA ; Geobacillus thermoleovorans ¦Á-amylase ; gDNA ; genomic DNA ; ThMA ; Thermus maltogenic amylase ; CGTase ; cyclodextrin glycosyltransferase ; TVAII ; Thermoactinomyces vulgaris R-47 ¦Á-amylase 2 ; SBD ; starch-binding domain ; AMY1 ; barley ¦Á-amylase isozyme 1
• 刊名：Enzyme and Microbial Technology
• 出版年：2013
• 出版时间：10 June, 2013
• 年：2013
• 卷：53
• 期：1
• 页码：46-54
• 全文大小：4317 K

文摘

A truncated form of an ¦Á-amylase, GTA, from thermophilic Geobacillus thermoleovorans CCB_US3_UF5 was biochemically and structurally characterized. The recombinant GTA, which lacked both the N- and C-terminal transmembrane regions, functioned optimally at 70 ¡ãC and pH 6.0. While enzyme activity was not enhanced by the addition of CaCl₂, GTA's thermostability was significantly improved in the presence of CaCl₂. The structure, in complex with an acarbose-derived pseudo-hexasaccharide, consists of the typical three domains and binds one Ca²⁺ ion. This Ca²⁺ ion was strongly bound and not chelated by EDTA. A predicted second Ca²⁺-binding site, however, was disordered. With limited subsites, two novel substrate-binding residues, Y147 and Y182, may help increase substrate affinity. No distinct starch-binding domain is present, although two regions rich in aromatic residues have been observed. GTA, with a smaller domain B and several shorter loops compared to other ¦Á-amylases, has one of the most compact ¦Á-amylase folds that may contribute greatly to its tight Ca²⁺ binding and thermostability.