Wulff Nelson Arno, Carrer Helaine, Pascholati Sérgio Florentino
Escola Superior de Agricultura "Luiz de Queiroz," USP, Departamento de Entomologia, Fitopatologia e Zoologia Agrícola, Av. Pádua Dias 11, CEP 13418-900, Piracicaba, SP, Brasil.
Curr Microbiol. 2006 Sep;53(3):198-203. doi: 10.1007/s00284-005-0475-2. Epub 2006 Jul 27.
Xylella fastidiosa was the first plant pathogen whose complete genome sequence was available. X. fastidiosa causes citrus variegated chlorosis, but the physiological basis of the disease in unknown. Through comparative sequence analysis, several putative plant cell wall-degrading enzymes were identified on the X. fastidiosa genome. We have cloned Xf818, a putative endoglucanase ORF, into expression vectors pET20b and pET28b, and purified a recombinant form of Xf818 containing a His(6) tag. Through biochemical assays, we have characterized the endoglucanase activity of this protein. The best conditions for hydrolysis over carboxymethyl cellulose (CMC) were on pH 5.2 at 65 degrees C. Xf818 hydrolyzed CMC, acid swollen cellulose, Avicel, birch wood, oat spels xylans, and the oligosaccharides cellotetraose and cellopentaose. Xf818 carried out transglycosylation and had a functional cellulose-binding domain.
木质部难养菌是首个全基因组序列已知的植物病原体。木质部难养菌会引发柑橘杂色黄化病，但其致病的生理基础尚不清楚。通过比较序列分析，在木质部难养菌基因组上鉴定出了几种假定的植物细胞壁降解酶。我们已将一个假定的内切葡聚糖酶开放阅读框Xf818克隆到表达载体pET20b和pET28b中，并纯化了一种带有His(6)标签的重组Xf818。通过生化分析，我们对该蛋白的内切葡聚糖酶活性进行了表征。水解羧甲基纤维素（CMC）的最佳条件是在pH 5.2、65℃。Xf818能水解CMC、酸溶胀纤维素、微晶纤维素、桦木、燕麦木聚糖以及纤维四糖和纤维五糖等寡糖。Xf818具有转糖基化作用且有一个功能性纤维素结合结构域。
