Mutational analysis of the N-terminal domain of UreR, the positive transcriptional regulator of urease gene expression.
Mutational analysis of the N-terminal domain of UreR, the positive transcriptional regulator of urease gene expression.
复制标题
DOI：
10.1016/j.micres.2012.03.005
复制
发表时间：
2012-07-25
影响因子：
6.7
通讯作者：
Collins, Carleen M.
中科院分区：
文献类型：
--
作者：
Parra, Maria C.;Collins, Carleen M.
大肠杆菌质粒编码的尿素酶是人类和动物泌尿道和胃十二指肠感染中的毒力因子，当底物尿素存在于生长培养基中时会被诱导。尿素依赖性脲酶的表达在转录水平上由AraC样激活剂UreR介导。先前的工作表明，代表UreR的N-末端194个氨基酸残基的肽在单个位点结合尿素，全长UreR形成寡聚体，并且寡聚基序被认为位于分子的N-末端部分。UreR的C末端结构域包含两个螺旋-转角-螺旋基序，推测这是DNA结合所必需的。在这项研究中，我们利用突变分析的N-末端结构域的UreR，以确定该域的二聚类似于其他AraC家族成员。分析UreR突变体激活来自ureDp-lacZ转录融合体的lacZ转录的能力。编码UreR的N-末端194个氨基酸的构建体，通过凝胶过滤作为寡聚体洗脱，并且相对于野生型ureR等位基因具有显性阴性表型。我们推测，这种显性负性表型的野生型和截短的UreR之间的非活性异二聚体的形成的结果。显性阴性分析和交联试验证明E. coli UreR作为二聚体具有活性，并且二聚化发生在前180个残基内。
The Escherichia coli plasmid-encoded urease, a virulence factor in human and animal infections of the urinary and gastroduodenal tracts, is induced when the substrate urea is present in the growth medium. Urea-dependent urease expression is mediated at the transcriptional level by the AraC-like activator UreR. Previous work has shown that a peptide representing the N-terminal 194 amino-acid residues of UreR binds urea at a single site, full-length UreR forms an oligomer, and the oligomerization motif is thought to reside in the N-terminal portion of the molecule. The C-terminal domain of UreR contains two helix-turn-helix motifs presumed to be necessary for DNA binding. In this study, we exploited mutational analyses at the N-terminal domain of UreR to determine if this domain dimerizes similar to other AraC family members. UreR mutants were analyzed for the ability to activate transcription of lacZ from an ureDp-lacZ transcriptional fusion. A construct encoding the N-terminal 194 amino acids of UreR, eluted as an oligomer by gel filtration and had a dominant negative phenotype over the wild-type ureR allele. We hypothesize that this dominant negative phenotype results from the formation of inactive heterodimers between wild-type and truncated UreR. Dominant negative analysis and cross-linking assays demonstrated that E. coli UreR is active as a dimer and dimerization occurs within the first 180 residues.
登录
查看更多内容
DOI：
10.1073/pnas.90.12.5638
发表时间：
1993-06-15
影响因子：
11.1
作者：
BUSTOS, SA;SCHLEIF, RF
通讯作者：
SCHLEIF, RF
影响因子：
3.2
作者：
Dirla, Stephanie;Chien, John Yeh-Heng;Schleif, Robert
通讯作者：
Schleif, Robert
影响因子：
3.2
作者：
MULROONEY, SB;LYNCH, MJ;HAUSINGER, RP
通讯作者：
HAUSINGER, RP
影响因子：
3.6
作者：
Paytubi, S;Madrid, C;Juárez, A
通讯作者：
Juárez, A
影响因子：
5.6
作者：
Childers, Brandon M.;Weber, Gregor G.;Klose, Karl E.
通讯作者：
Klose, Karl E.