Feng P, Lampel K A
Division of Microbiological Studies, Food and Drug Administration, Washington, DC 20204.
Microbiology (Reading). 1994 Aug;140 ( Pt 8):2101-7. doi: 10.1099/13500872-140-8-2101.
Isolates of enterohaemorrhagic Escherichia coli serotype O157:H7 do not exhibit beta-glucuronidase (GUD) activity; however, they carry nucleotide sequences for the uidA gene that encodes the GUD enzyme. Polymerase chain reaction analysis using uidA-specific primers confirmed that a genetic region homologous in size to the E. coli uidA structural gene, including the regulatory region, was present in E. coli O157:H7 isolates. DNA sequencing analysis of the regulatory region and the 5' terminus of the uidA gene of E. coli O157:H7 showed a base substitution in the putative -10 promotor region and at 93 bases downstream from the initiation codon. Neither base alteration, however, appeared to affect uidA allele gene expression in the O157:H7 isolates. Immunoblotting of cell extracts with an anti-GUD antibody showed that E. coli O157:H7 isolates produced an antibody-reactive protein that was homologous in size to E. coli GUD, but no GUD activity was observed in cell-free extracts of these isolates. These results suggest that the antibody-reactive protein produced by E. coli O157:H7 may be an inactive GUD enzyme. Sequencing of the uidA structural gene in O157:H7 showed the presence of 18 additional nucleotide base substitutions, but only six altered the amino acid sequence. Also, there were two frame shift mutations, 18 bases apart, that altered the sequence of six consecutive amino acids. These genetic alterations in the uidA structural gene of O157:H7 may account for the absence of GUD activity in this serotype.
肠出血性大肠杆菌O157:H7血清型菌株不表现出β-葡萄糖醛酸酶（GUD）活性；然而，它们携带编码GUD酶的uidA基因的核苷酸序列。使用uidA特异性引物进行的聚合酶链反应分析证实，在大肠杆菌O157:H7菌株中存在一个与大肠杆菌uidA结构基因大小同源的遗传区域，包括调控区域。对大肠杆菌O157:H7的调控区域和uidA基因5'末端的DNA测序分析显示，在假定的-10启动子区域以及起始密码子下游93个碱基处存在碱基替换。然而，这两种碱基改变似乎都不影响O157:H7菌株中uidA等位基因的表达。用抗GUD抗体对细胞提取物进行免疫印迹分析表明，大肠杆菌O157:H7菌株产生了一种与大肠杆菌GUD大小同源的抗体反应性蛋白，但在这些菌株的无细胞提取物中未观察到GUD活性。这些结果表明，大肠杆菌O157:H7产生的抗体反应性蛋白可能是一种无活性的GUD酶。对O157:H7中uidA结构基因的测序显示，还存在另外18个核苷酸碱基替换，但只有6个改变了氨基酸序列。此外，还有两个移码突变，相隔18个碱基，改变了六个连续氨基酸的序列。O157:H7的uidA结构基因中的这些遗传改变可能是该血清型缺乏GUD活性的原因。
