Glycosylation and Biogenesis of Streptococcal Adhesins

链球菌粘附素的糖基化和生物合成

• 批准号: 10227893
• 负责人: Hui Wu
• 金额: $ 36.29万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-03 至 2023-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10227893
• 关键词: Bacteria; Bacterial Adhesins; Bacterial Infections; Bacterial Proteins; Basic Science; Big Data; Biochemical; Biochemistry; Biogenesis; Biological Process; Biology; Complex; Crystallization; Development; Devices; Elements; Endoplasmic Reticulum; Environment; Enzymes; Escherichia coli; Exhibits; Family; Funding; Future; GTPase-Activating Proteins; Gene Cluster; Genetic; Genomics; Glucosyltransferase; Glycobiology; Glycoproteins; Glycoside Hydrolases; Gram-Positive Bacteria; Homeostasis; Interdisciplinary Study; Lectin; Link; Mediating; Microbial Biofilms; Modeling; Molecular; Molecular Chaperones; Names; Oral; Oral Microbiology; Oral cavity; Oral health; Organism; Pathway interactions; Peer Review; Phenotype; Play; Polysaccharides; Process; Protein Family; Protein Glycosylation; Protein Precursors; Protein Secretion; Proteins; Proteomics; Publishing; Quality Control; Recombinants; Reporting; Research; Resolution; Role; Series; Serine; Staphylococcus aureus; Streptococcus; Streptococcus adhesin; Structure; System; Testing; Virulence; Work; bacterial community; bacterial fitness; combat; frontier; glucosidase; glycosylation; glycosyltransferase; in vivo; insight; microbial; microbiota; novel; novel strategies; oral biofilm; oral commensal; oral streptococci; pathogen; pathogenic bacteria; protein complex; protein folding; structural biology; therapeutic target; three dimensional structure

Essential to oral biofilm development is the initial colonization by oral streptococci. The abundant oral streptococci keep pathogens at bay. We have used the most abundant oral streptococcus, Streptococcus parasanguinis as a model to study bacterial colonization and identified a new family of bacterial serine-rich repeat proteins (SRRPs) named “fimbriae-associated protein-1” (Fap1). Fap1 is heavily glycosylated, and glycosylation of Fap1 is crucial for bacterial biofilm formation. Since our discovery of Fap1, Fap1-like SRRPs have been identified from numerous Gram-positive bacteria and implicated in bacterial fitness and virulence. Our studies have led to the groundbreaking discovery of a new Fap1 biosynthetic pathway. We have demonstrated that the Fap1 biogenesis is controlled by a gene cluster encoding a series of novel glycosyltransferases and unique accessory secretion proteins. Biogenesis of SRRPs has now emerged as a new paradigm to investigate bacterial protein glycosylation and secretion. During our study of Fap1 glycosylation, we have defined a complete glycosylation pathway that synthesizes a novel Fap1 glycan. In the study of Fap1 secretion, we have identified a protein complex consisting of three distinct glycosylation associated proteins Gap1, 2 and 3 that work in concert to modulate the Fap1 maturation and biogenesis. Further, we have determined the high resolution 3-dimensional structure of the Gap1/2/3 complex, which uncovered new mechanistic insights for this 3-protein complex. Gap1 and Gap2 exhibit dual functions in the biogenesis of Fap1. 1), Gap1 and Gap2 modulates the formation of the protein complex as a molecular chaperone. 2), Gap1 and Gap2 function as a glucosyltransferase and glucosidase respectively in the quality control of Fap1 maturation and biogenesis. The Gap protein complex resembles the three-key elements in the eukaryotic quality control system dedicated to glycosylated proteins, hence we will continue our basic science discovery of new biology and biochemistry linked to maturation and biogenesis of Fap1 & other SRRPs. Aim 1 Determine how Gap1 functions as a molecular chaperone for Gap2 and as a key quality control glycosyltransferase to process Fap1 precursor during Fap1 biogenesis. We will use genetic, biochemical, structural biology, and glycobiology approaches to investigate how Gap1 stabilizes Gap2 as a molecular chaperone, and how Gap1 acts as a quality control glycosyltransferase to process Fap1 precursor. Aim 2 Define the roles played by Gap2 as a molecular chaperone for Gap3 and as a key quality control glucosidase during Fap1 biogenesis. We will determine how Gap2 assists Gap3 as an accessory chaperone, and coordinates with Gap1 and Gap3 to process Fap1, thus modulating the Fap1 biogenesis. As biogenesis of SRRPs is highly conserved in Gram-positive bacteria, deciphering novel molecular insight to this new protein complex as a quality control system will offer new opportunities to develop new strategies to maintain healthy oral cavity as well as to combat bacterial infections.

口腔生物被膜的形成主要是口腔链球菌的初始定植。丰富的口腔 链球菌将病原体拒之门外我们使用了最丰富的口腔链球菌， parasanguinis作为研究细菌定植的模型，并确定了一个新的富含丝氨酸的细菌家族， 重复蛋白（SRRP），命名为“菌毛相关蛋白-1”（Fap 1）。Fap 1是高度糖基化的，并且 Fap 1的糖基化对于细菌生物膜的形成至关重要。自从我们发现了Fap 1，Fap 1样SRRP 已从许多革兰氏阳性细菌中鉴定出，并与细菌适应性和毒力有关。 我们的研究导致了一个新的Fap 1生物合成途径的突破性发现。我们有 证明Fap 1的生物发生是由一个基因簇控制的，该基因簇编码一系列新的 糖基转移酶和独特的辅助分泌蛋白。SRRP的生物发生现在已经成为一种 研究细菌蛋白质糖基化和分泌的新范式。在我们对Fap 1的研究中， 通过糖基化，我们已经定义了合成新型Fap 1聚糖的完整糖基化途径。在 通过对Fap 1分泌的研究，我们鉴定了由三种不同糖基化组成的蛋白质复合物 相关蛋白Gap 1、2和3协同作用以调节Fap 1成熟和生物发生。 此外，我们还确定了Gap 1/2/3复合物的高分辨率三维结构， 发现了这个3-蛋白质复合物的新机制。Gap 1和Gap 2在细胞中具有双重功能， Fap 1的生物起源1），Gap 1和Gap 2调节蛋白质复合物作为分子的形成 监护人2）Gap 1和Gap 2分别作为葡萄糖基转移酶和葡萄糖苷酶在品质中发挥作用 控制Fap 1成熟和生物发生。差距蛋白复合物类似于三个关键要素， 真核生物质量控制系统致力于糖基化蛋白质，因此，我们将继续我们的基础科学 发现与Fap 1和其他SRRP的成熟和生物发生相关的新生物学和生物化学。 目的1确定Gap 1如何作为Gap 2的分子伴侣和关键质量控制 糖基转移酶在Fap 1生物合成过程中加工Fap 1前体。我们将使用遗传学，生物化学， 结构生物学和糖生物学方法来研究Gap 1如何稳定Gap 2作为一种分子， 分子伴侣，以及Gap 1如何作为质量控制糖基转移酶来加工Fap 1前体。 目的2明确Gap 2作为Gap 3的分子伴侣和关键质量控制的作用 Fap 1生物发生过程中的葡萄糖苷酶。我们将确定Gap 2如何协助Gap 3作为一个附件 分子伴侣，并与Gap 1和Gap 3协调以加工Fap 1，从而调节Fap 1生物合成。 由于SRRP的生物发生在革兰氏阳性菌中高度保守，因此可以解释新的分子见解， 这种新的蛋白质复合物作为质量控制系统将为开发新的策略提供新的机会， 保持口腔健康，对抗细菌感染。