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.

口服生物膜发育至关重要的是口服链球菌的初始定植。丰富的口服 链球菌使病原体陷入困境。我们使用了最丰富的口腔链球菌，链球菌 副parasanguinis是研究细菌定植并鉴定出新的细菌丝氨酸家族的模型 重复蛋白（SRRP）称为“ fimbriae相关蛋白-1”（FAP1）。 FAP1是大量糖基化的，并且 FAP1的糖基化对于细菌生物膜形成至关重要。自从我们发现FAP1，类似FAP1的SRRP以来 已经从许多革兰氏阳性细菌中鉴定出来，并暗示了细菌适应性和病毒。 我们的研究导致了新的FAP1生物合成途径的开创性发现。我们有 证明FAP1生物发生由编码一系列新颖的基因簇控制 糖基转移酶和独特的附件分泌蛋白。 SRRP的生物发生现已成为 研究细菌蛋白糖基化和分泌的新范式。在我们研究FAP1期间 糖基化，我们定义了一个完整的糖基化途径，该途径合成了一种新型的FAP1聚糖。在 FAP1分泌的研究，我们确定了由三种不同的糖基化组成的蛋白质复合物 相关的蛋白质GAP1、2和3共同调节FAP1成熟和生物发生。 此外，我们已经确定了GAP1/2/3复合物的高分辨率3维结构，该结构 发现了这种三蛋白复合物的新机械见解。 GAP1和GAP2在 FAP1的生物发生。 1），GAP1和GAP2调节蛋白质复合物的形成为分子 伴侣。 2），GAP1和GAP2分别充当葡萄糖基转移酶和葡萄糖酶的质量 控制FAP1成熟和生物发生。 GAP蛋白复合物类似于 真核质量控制系统专用于糖基化蛋白，因此我们将继续我们的基础科学 发现与FAP1和其他SRRP的成熟和生物发生有关的新生物学和生物化学。 AIM 1确定GAP1如何用作GAP2的分子伴侣和关键质量控制 糖基转移酶在FAP1生物发生过程中处理FAP1前体。我们将使用遗传，生化， 结构生物学和糖生物学方法，用于研究GAP1如何稳定GAP2作为分子 伴侣，以及GAP1如何充当质量控制糖基转移酶来处理FAP1前体。 AIM 2将GAP2扮演的角色定义为GAP3的分子伴侣和关键质量控制 FAP1生物发生过程中的葡萄糖酶。我们将确定GAP2如何帮助GAP3作为配件 伴侣，并与GAP1和GAP3坐标以处理FAP1，从而调节FAP1生物发生。 由于SRRP的生物发生在革兰氏阳性细菌中高度保守，因此将新颖的分子见解解密到 这个新的蛋白质复合物作为质量控制系统将为制定新策略提供新的机会 保持健康的口腔以及对抗细菌感染。