Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, MD 21201-1509, USA.
J Bacteriol. 2011 Feb;193(4):875-86. doi: 10.1128/JB.00922-10. Epub 2010 Dec 10.
Enterohemorrhagic Escherichia coli (EHEC) is a food-borne pathogen that can cause severe health complications and utilizes a much lower infectious dose than other E. coli pathotypes. Despite having an intact ure locus, ureDABCEFG, the majority of EHEC strains are phenotypically urease negative under tested conditions. Urease activity potentially assists with survival fitness by enhancing acid tolerance during passage through the stomach or by aiding with colonization in either human or animal reservoirs. Previously, in the EHEC O157:H7 Sakai strain, a point mutation in ureD, encoding a urease chaperone protein, was identified, resulting in a substitution of an amber stop codon for glutamine. This single nucleotide polymorphism (SNP) is observed in the majority of EHEC O157:H7 isolates and correlates with a negative urease phenotype in vitro. We demonstrate that the lack of urease activity in vitro is not solely due to the amber codon in ureD. Our analysis has identified two additional SNPs in ureD affecting amino acid positions 38 and 205, in both cases determining whether the encoded amino acid is leucine or proline. Phylogenetic analysis based on Ure protein sequences from a variety of urease-encoding bacteria demonstrates that the proline at position 38 is highly conserved among Gram-negative bacteria. Experiments reveal that the L38P substitution enhances urease enzyme activity; however, the L205P substitution does not. Multilocus sequence typing analysis for a variety of Shiga toxin-producing E. coli isolates combined with the ureD sequence reveals that except for a subset of the O157:H7 strains, neither the in vitro urease-positive phenotype nor the ureD sequence is phylogenetically restricted.
肠出血性大肠杆菌（EHEC）是一种食源性病原体，可引起严重的健康并发症，其感染剂量比其他大肠杆菌血清型低得多。尽管具有完整的 ure 基因座 ureDABCEFG，但在测试条件下，大多数 EHEC 菌株的表型呈尿素酶阴性。尿素酶活性可能通过增强在胃中通过时的耐酸性，或通过有助于在人类或动物储层中定植，从而有助于生存适应性。先前，在 EHEC O157:H7 Sakai 菌株中，鉴定出 ureD 编码尿素酶伴侣蛋白的点突变，导致谷氨酰胺取代琥珀终止密码子。这种单核苷酸多态性（SNP）在大多数 EHEC O157:H7 分离株中观察到，并与体外尿素酶阴性表型相关。我们证明体外缺乏尿素酶活性不仅是由于 ureD 中的琥珀密码子。我们的分析还确定了 ureD 中另外两个影响氨基酸位置 38 和 205 的 SNP，在这两种情况下，决定了编码的氨基酸是亮氨酸还是脯氨酸。基于来自各种具有尿素酶编码基因的细菌的 ure 蛋白序列的系统发育分析表明，位置 38 的脯氨酸在革兰氏阴性菌中高度保守。实验表明，L38P 取代增强了尿素酶酶活性；然而，L205P 取代则没有。对各种产志贺毒素大肠杆菌分离株的多位点序列分型分析结合 ureD 序列表明，除了一部分 O157:H7 菌株外，体外尿素酶阳性表型和 ureD 序列在系统发育上都不受限制。
