ROLE OF SURFACE MOLECULES IN COLONIZATION AND BIOLOGICAL MIMICRY

表面分子在定植和生物拟态中的作用

• 批准号: 5201785
• 负责人: J LONDON
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF DENTAL & CRANIOFACIAL RESEARCH
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/5201785
• 关键词: Escherichia coli; adhesin; bacterial genetics; bacterial proteins; beta galactosidase; ecology; enzyme activity; enzyme biosynthesis; genetic translation; host organism interaction; membrane proteins; messenger RNA; molecular cloning; nucleic acid probes; nucleic acid sequence; nucleotides; oral bacteria; pilus; protein sequence; site directed mutagenesis

Bacterial colonization and invasion depend upon a microbe's ability to attach to a host and multiply. Oral bacteria, like other human commensals and pathogens, have evolved specific surface proteins for this purpose. Some of these molecules are associated with scaffolding structures, e.g., pili or fimbriae, and others are integral parts of the organism's outer membrane. Several adhesins on the surface of Prevotella loescheii have been the focus of studies in this laboratory. The gene for a 75 kDa lectin-like protein was cloned and its structure deduced from the nucleotide sequence. Translation of the adhesin mRNA occurs via a frameshifting hop in which 27 nucleotides are bypassed. Since bypasses are rarely seen among prokaryotes, it was essential to establish that the amino acid sequences prior to and immediately following the hop were the expected residues. Recent attempts to sequence this region yielded the anticipated amino acid sequence proving that a 27 nucleotide bypass occurs during translation. Elements of the "hop", e.g., the shift region, stem loop and pseudoknot are being assessed for their effect on translation of the adhesin and heterologous proteins. Beta-galactosidase constructs with the 228 nucleotide hop positioned in frame at the 5' end of the gene indicated that synthesis of the enzyme in Escherichia coli was being retarded compared to wild type production. Current studies using site-specific mutagenesis have demonstrated that regions within the stem loop and pseudoknot are absolutely essential for read-through of the adhesin mRNA. Attempts to identify and clone the gene encoding a P. loescheii non-lectin adhesin using degenerated probes based on the first 10 amino acids of the N-terminal sequence are continuing. The gene of a P. loescheii surface component believed to be a support protein was cloned and sequenced. Comparisons indicated that the deduced sequence showed extensive identity to both eukaryotic and prokaryotic enolases. The partially purified protein from which the N-terminal amino acid sequence was obtained shows enolase activity. The role of this enzyme is being investigated.

细菌定植和入侵取决于微生物的能力， 附着在宿主上繁殖 口腔细菌，像其他人类口腔细菌一样， 和病原体，已经进化出用于此目的的特异性表面蛋白。 这些分子中的一些与支架结构相关，例如， 皮利或菌毛等是生物体外膜的组成部分， 膜的 罗氏普雷沃菌表面的几种粘附素 是这个实验室研究的重点。 75 kDa的基因 克隆了凝集素样蛋白，并从 核苷酸序列。 粘附素mRNA的翻译是通过一个 移码跳跃，其中27个核苷酸被绕过。 由于旁路 在原核生物中很少见，因此必须确定， 在啤酒花之前和之后的氨基酸序列是 预期残留物。 最近尝试对该区域进行测序， 预期的氨基酸序列证明发生了27个核苷酸的旁路 在翻译过程中。 “跳”的元素，例如，移位区、茎 正在评估环和假结对平移的影响， 粘附素和异源蛋白。 β-半乳糖苷酶构建体， 该228个核苷酸跳跃位于该基因5’端的读码框内 这表明，大肠杆菌中的酶的合成是 与野生型生产相比延迟。 目前的研究使用 定点诱变已经证明茎内的区域 循环和伪结对于通读 粘附素mRNA。 试图鉴定和克隆编码P. 使用基于第一种方法的简并探针的洛氏非凝集素粘附素 N末端序列的10个氨基酸是连续的。 A的基因 P.克隆了被认为是支持蛋白的Loescheii表面组分 和序列测定 比较表明，推导的序列显示， 与真核和原核烯醇化酶具有广泛的同一性。 的 部分纯化的蛋白质，其中N末端氨基酸序列 显示烯醇化酶活性。 这种酶的作用是 研究了