Genetic characterization of wild-type and mutant fur genes of Bordetella avium.
Genetic characterization of wild-type and mutant fur genes of Bordetella avium.
复制标题
鸟博德特氏菌野生型和突变型毛皮基因的遗传特征。
DOI：
10.1128/iai.67.6.3160-3165.1999
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发表时间：
1999
影响因子：
3.1
通讯作者：
Connell,TD
中科院分区：
文献类型：
--
作者：
Murphy,ER;Dickenson,A;Militello,KT;Connell,TD
对于大多数(如果不是全部)生物体来说，铁(Fe)是一种基本元素。为了应对铁的营养需求，细菌进化出复杂的系统，从环境中获取铁元素。编码这些系统的基因通常是协同调节的，以响应铁的浓度。最近的研究表明，禽类呼吸道病原体Bordeella avium表达许多铁调节基因(T.D.Connell、A.Dickenson、A.J.Marone、K.T.Militello、M.J.Filiatraut、M.L.Hayman和J.Pitula)。伊蒙。66：3597-3605,1998)。对一株工程菌株OB进行了锰的筛选。携带铁调节的碱性磷酸酶报告基因，获得了一个铁调节基因表达受影响的突变体。以确定铁依赖的调节是否在INB。Avium是由theb的一个片段--类afur基因介导的。Avium染色体，与B.用聚合酶链式反应(PCR)克隆了百日咳病毒。测序结果表明，该片段来自B。Avium编码的多肽与毛皮蛋白OFB有92%的同源性。百日咳。体内实验表明，克隆的基因与大肠杆菌突变株H1780互补。Southern杂交和PCRS分析表明，该突变株在呋喃烯阅读框的5‘端有2~3kbp的核苷酸序列缺失。这是一种自发的由聚合酶链式反应产生的突变体。该基因编码18位氨基酸(R18H)上的组氨酸取代精氨酸的Fur蛋白。遗传分析表明，R18H突变基因克隆到低拷贝数载体后不能补充H1780中的突变。然而，当将R18H突变基因克隆到高拷贝数的载体上时，该突变基因能够补充突变。克隆的野生型呋喃基因将作为一种遗传工具用于鉴定THb中毛皮调节基因。鸟巢染色体。
For most, if not all, organisms, iron (Fe) is an essential element. In response to the nutritional requirement for Fe, bacteria evolved complex systems to acquire the element from the environment. The genes encoding these systems are often coordinately regulated in response to the Fe concentration. Recent investigations revealed thatBordetella avium, a respiratory pathogen of birds, expressed a number of Fe-regulated genes (T. D. Connell, A. Dickenson, A. J. Martone, K. T. Militello, M. J. Filiatraut, M. L. Hayman, and J. Pitula, Infect. Immun. 66:3597–3605, 1998). By using manganese selection on an engineered strain ofB. aviumthat carried an Fe-regulated alkaline phosphatase reporter gene, a mutant was obtained that was affected in expression of Fe-regulated genes. To determine if Fe-dependent regulation inB. aviumwas mediated by afur-like gene, a fragment of theB. aviumchromosome, corresponding to thefurlocus ofB. pertussis, was cloned by PCR. Sequencing revealed that the fragment fromB. aviumencoded a polypeptide with 92% identity to the Fur protein ofB. pertussis. In vivo experiments showed that the cloned gene complemented H1780, afurmutant ofEscherichia coli. Southern hybridizations and PCRs demonstrated that the manganese mutant had a deletion of 2 to 3 kbp of nucleotide sequence in the region located immediately 5′ of thefuropen reading frame. A spontaneous PCR-derived mutant of theB. avium furgene was isolated that encoded a Fur protein in which a histidine was substituted for an arginine at amino acid position 18 (R18H). Genetic analysis showed that the R18H mutant gene when cloned into a low-copy-number vector did not complement thefurmutation in H1780. However, the R18H mutant gene was able to complement thefurmutation when cloned into a high-copy-number vector. The cloned wild-typefurgene will be useful as a genetic tool to identify Fur-regulated genes in theB. aviumchromosome.