Iyo A H, Forsberg C W
Department of Microbiology, University of Guelph, ON, Canada.
Can J Microbiol. 1996 Sep;42(9):934-43. doi: 10.1139/m96-120.
A 3.6-kb fragment of the Fibrobacter succinogenes S85 DNA was sequenced and found to contain two open reading frames (ORFs) on the same strand separated by 242 nucleotide bases. The translated protein from ORF1 had a predicted mass of 52.3 kDa. In a region of 320 amino acid overlap, it shares a 35% identity with the b-chain of the glutamate synthase of Escherichia coli. The ORF2 protein encodes a 519 residue protein designated CelG. It consists of an ORF of 1557 bp, encoding a polypeptide of 54.5 kDa. The N-terminal region, which contains the catalytic domain, is linked to a C-terminal basic domain, which has a predicted isoelectric point of 10.8. The catalytic domain in endoglucanase G (CelG) is homologous to the family 5 (A) cellulases. The enzyme has an apparent mass of 55 kDa, a pH optimum of 5.5, and temperature optimum of 25 degrees C. It had a specific activity of 16.5 mumols x min(-1) x mg-1 on barley b-glucan and produced a mixture of cellooligosaccharides from the hydrolysis of acid swollen cellulose and cellooligosaccharides. Antiserum raised against the purified form of CelG in E. coli failed to react with proteins from the native organism when grown on either glucose or crystalline cellulose, but reverse transcription and polymerase chain reaction techniques using RNA from the native organism demonstrated that the celG gene was expressed constitutively. Its distribution amongst subspecies of Fibrobacter was restricted to F. succinogenes S85.
对琥珀酸纤维杆菌S85的一段3.6 kb DNA片段进行了测序，发现该片段在同一条链上含有两个开放阅读框（ORF），它们被242个核苷酸碱基隔开。来自ORF1的翻译后蛋白质预测分子量为52.3 kDa。在320个氨基酸的重叠区域，它与大肠杆菌谷氨酸合酶的b链具有35%的同一性。ORF2蛋白编码一个由519个残基组成的蛋白质，命名为CelG。它由一个1557 bp的ORF组成，编码一个54.5 kDa的多肽。包含催化结构域的N端区域与C端碱性结构域相连，其预测的等电点为10.8。内切葡聚糖酶G（CelG）中的催化结构域与第5（A）家族纤维素酶同源。该酶的表观分子量为55 kDa，最适pH为5.5，最适温度为25℃。它对大麦β-葡聚糖的比活性为16.5 μmol·min⁻¹·mg⁻¹，能将酸膨胀纤维素水解产生纤维寡糖混合物。用在大肠杆菌中纯化的CelG制备的抗血清，在天然菌株以葡萄糖或结晶纤维素为碳源生长时，不能与天然菌株的蛋白质发生反应，但利用天然菌株的RNA进行逆转录和聚合酶链反应技术表明celG基因是组成型表达的。它在纤维杆菌亚种中的分布仅限于琥珀酸纤维杆菌S85。
