Yaoi Katsuro, Mitsuishi Yasushi
Institute for Biological Resources and Functions, National Institute of Advanced Industrial Science and Technology, Tsukuba Central 6, 1-1-1 Higashi, Ibaraki 305-8566, Japan.
J Biol Chem. 2002 Dec 13;277(50):48276-81. doi: 10.1074/jbc.M208443200. Epub 2002 Oct 8.
A novel oligoxyloglucan-specific glycosidase, oligoxyloglucan reducing end-specific cellobiohydrolase (OXG-RCBH), with a molecular mass of 97 kDa and a pI of 6.1, was isolated from the fungus Geotrichum sp. M128. Analysis of substrate specificity using various xyloglucan oligosaccharide structures revealed that OXG-RCBH had exoglucanase activity. It recognized the reducing end of oligoxyloglucan and released two glucosyl residue segments from the main chain. The full-length cDNA encoding OXG-RCBH was cloned and sequenced, and it had a 2436-bp open reading frame encoding an 812amino acid protein. The deduced protein showed approximately 35% identity to members of glycoside hydrolase family 74. The cDNA encoding OXG-RCBH was then expressed in Escherichia coli. Although the recombinant protein was expressed as an inclusion body, renaturation was successful, and enzymatically active recombinant OXG-RCBH was obtained.
从真菌地霉属M128中分离出一种新型的低聚木葡聚糖特异性糖苷酶，即低聚木葡聚糖还原端特异性纤维二糖水解酶（OXG-RCBH），其分子量为97 kDa，等电点为6.1。使用各种木葡聚糖寡糖结构分析底物特异性表明，OXG-RCBH具有外切葡聚糖酶活性。它识别低聚木葡聚糖的还原端，并从主链上释放出两个葡萄糖基残基片段。克隆并测序了编码OXG-RCBH的全长cDNA，其具有一个2436 bp的开放阅读框，编码一个812个氨基酸的蛋白质。推导的蛋白质与糖苷水解酶家族74的成员显示出约35%的同一性。然后将编码OXG-RCBH的cDNA在大肠杆菌中表达。尽管重组蛋白以包涵体形式表达，但复性成功，获得了具有酶活性的重组OXG-RCBH。
