Liu Y N, Tang J L, Clarke B R, Dow J M, Daniels M J
Sainsbury Laboratory, John Innes Institute, Norwich, UK.
Mol Gen Genet. 1990 Feb;220(3):433-40. doi: 10.1007/BF00391750.
A multipurpose broad host range plasmid, pIJ3200, was constructed by inserting the polylinker-containing 445 bp PvuII fragment of Bluescript M13 into the EcoRI site of the cosmid pLAFR1. Using this vector a protease gene of Xanthomonas campestris pathovar campestris, previously cloned in the recombinant plasmid pIJ3070, was located by deletion to a 2.2 kb DNA region. Sequencing of the protease gene revealed an open reading frame encoding a 580 amino acid polypeptide with molecular weight of 57,000. The deduced amino acid sequence showed strong homology with the subtilisin family of serine proteases. This, together with its sensitivity to inhibition by phenylmethylsulphonyl fluoride, suggests that the enzyme belongs to this family of proteases.
通过将含多克隆位点的445 bp Bluescript M13 PvuII片段插入黏粒pLAFR1的EcoRI位点，构建了一个多用途广宿主范围质粒pIJ3200。利用该载体，通过缺失定位，将先前克隆于重组质粒pIJ3070中的野油菜黄单胞菌野油菜致病变种的一个蛋白酶基因定位于一个2.2 kb的DNA区域。对该蛋白酶基因进行测序，发现一个编码580个氨基酸多肽、分子量为57,000的开放阅读框。推导的氨基酸序列与丝氨酸蛋白酶枯草杆菌蛋白酶家族具有高度同源性。这一点，再加上其对苯甲基磺酰氟抑制的敏感性，表明该酶属于这个蛋白酶家族。
