Goda Sayed K, Eissa Omima, Akhtar Muhammad, Minton Nigel P
Department of Molecular Microbiology, Centre for Applied Microbiology and Research, Porton Down Salisbury SP4 OJG, UK.
Department of Biochemistry, University of Southampton, Bassett Crescent East, Southampton SO 16 7PX, UK.
Microbiology (Reading). 1997 Oct;143 ( Pt 10):3287-3294. doi: 10.1099/00221287-143-10-3287.
An Escherichia coli clone was detected in a Clostridium butyricum NCIMB 7423 plasmid library capable of degrading soluble amylose. Deletion subcloning of its recombinant plasmid indicated that the gene(s) responsible for amylose degradation was localized on a 1.8 kb NspHI-Scal fragment. This region was sequenced in its entirety and shown to encompass a large ORF capable of encoding a protein with a calculated molecular mass of 57,184 Da. Although the deduced amino acid sequence showed only weak similarity with known amylases, significant sequences identity was apparent with the 4-alpha-glucano-transferase enzymes of Streptococcus pneumoniae (46.9%), potato (42.9%) and E. coli (16.2%). The clostridial gene (designated maIQ) was followed by a second ORF which, through its homology to the equivalent enzymes of E. coli and S. pneumoniae, was deduced to encode maltodextrin phosphorylase (MaIP). The translation stop codon of MaIQ overlapped the translation start codon of the putative maIP gene, suggesting that the two genes may be both transcriptionally and translationally coupled. 4-alpha-Glucanotransferase catalyses a disproportionation reaction in which single or multiple glucose units from oligosaccharides are transferred to the 4-hydroxyl group of acceptor sugars. Characterization of the recombinant C. butyricum enzyme demonstrated that glucose, maltose and maltotriose could act as acceptor, whereas of the three only maltotriose could act as donor. The enzyme therefore shares properties with the E. coli MaIQ protein, but differs significantly from the glucanotransferase of Thermotoga maritima, which is unable to use maltotriose as donor or glucose as acceptor. Physiologically, the concerted action of 4-alpha-glucanotransferase and maltodextrin phosphorylase provides C. butyricum with a mechanism of utilizing amylose/maltodextrins with little drain on cellular ATP reserves.
在丁酸梭菌NCIMB 7423质粒文库中检测到一个能够降解可溶性直链淀粉的大肠杆菌克隆。对其重组质粒进行缺失亚克隆表明，负责直链淀粉降解的基因位于一个1.8 kb的NspHI-Scal片段上。对该区域进行了全序列测定，结果显示它包含一个大的开放阅读框，能够编码一种计算分子量为57,184 Da的蛋白质。虽然推导的氨基酸序列与已知淀粉酶只有微弱的相似性，但与肺炎链球菌（46.9%）、马铃薯（42.9%）和大肠杆菌（16.2%）的4-α-葡聚糖转移酶有明显的序列同一性。梭菌基因（命名为maIQ）后面跟着第二个开放阅读框，通过与大肠杆菌和肺炎链球菌的等效酶的同源性推断，该开放阅读框编码麦芽糖糊精磷酸化酶（MaIP）。MaIQ的翻译终止密码子与假定的maIP基因的翻译起始密码子重叠，这表明这两个基因可能在转录和翻译上都是偶联的。4-α-葡聚糖转移酶催化一种歧化反应，其中寡糖中的单个或多个葡萄糖单位被转移到受体糖的4-羟基上。重组丁酸梭菌酶的特性表明，葡萄糖、麦芽糖和麦芽三糖可以作为受体，而在这三种糖中只有麦芽三糖可以作为供体。因此，该酶与大肠杆菌的MaIQ蛋白具有共同特性，但与嗜热栖热菌的葡聚糖转移酶有很大不同，嗜热栖热菌的葡聚糖转移酶不能使用麦芽三糖作为供体或葡萄糖作为受体。在生理上，4-α-葡聚糖转移酶和麦芽糖糊精磷酸化酶的协同作用为丁酸梭菌提供了一种利用直链淀粉/麦芽糖糊精的机制，而对细胞ATP储备的消耗很少。
