Structural Characterization of the TOC Protein Translocon Machinery

TOC 蛋白易位机的结构表征

• 批准号: 10376194
• 负责人: Nicholas Noinaj
• 金额: $ 31.54万
• 依托单位: PURDUE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-05-01 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10376194
• 关键词: Antibiotics; Apicomplexa; Artificial Membranes; Biological Assay; Biological Models; Biology; Cause of Death; Chloroplasts; Communicable Diseases; Complex; Country; Cryoelectron Microscopy; Cytoplasm; Defect; Destinations; Development; Drug usage; Energy Supply; Falciparum Malaria; Genes; Gram-Negative Bacteria; Guanosine Triphosphate Phosphohydrolases; Homology Modeling; Human; In Vitro; Infection; Malaria; Mediating; Membrane; Mitochondria; Modeling; Molecular; Molecular Chaperones; Molecular Machines; Mouse-ear Cress; Multi-Drug Resistance; Mutagenesis; N-terminal; Neurologic; Nuclear; Organelles; Parasites; Persons; Plasmodium falciparum; Plastids; Protein Export Pathway; Protein Import; Protein Precursors; Proteins; Reporting; Research; Resolution; Roentgen Rays; Role; Structure; System; Therapeutic; Toxoplasma; Toxoplasma gondii; Toxoplasmosis; X-Ray Crystallography; base; biophysical techniques; improved; nanodisk; novel; pathogen; polypeptide; protein transport; proteoliposomes; reconstitution; reconstruction; resistant strain; vaccine access

Project Summary Infectious diseases cause widespread sickness throughout the world each year and are the second leading cause of death, particularly in underdeveloped countries. And with the emergence of multi-drug resistance strains, the necessity for new, more effective, and more sustainable therapies is immediate. Included in these infectious diseases are the apicomplexa which includes Toxoplasma gondii and Plasmodium falciparum, the parasites causing toxoplasmosis and malaria, respectively. These parasites contain a unique plastid-like organelle called an apicoplast which contains four membranes and therefore have evolved a complex system for importing and exporting proteins across these membranes. These essential import/export machineries are ideal targets for novel antibiotics against these pathogens. Many of these translocon machineries are also conserved in other higher eukaryotic organelles such as chloroplasts and mitochondria, where a large majority of genes are nuclear encoded and therefore must be imported post-translationally. One such machinery is the conserved translocon of the outer membrane in chloroplasts (TOC) complex from Arabidopsis thaliana, a model system for studying chloroplast biology. The TOC complex consists of primarily three components, Toc33/34 and Toc159, both GTPases containing an N-terminal transmembrane helix anchoring them into the outer membrane, and Toc75, a 16- stranded β-barrel membrane-spanning translocon. While mechanistic models have been put forth for how the TOC complex functions, they have remained largely unproven due to the lack of structural characterization, which is needed to stitch together all the pieces of the mechanistic puzzle. In our studies, we will use biophysical methods, X-ray crystallography, cryo-electron microscopy, and small-angle X-ray scattering to structurally and functionally characterize this specialized machinery. Our results will fill a longstanding gap in the field and will be essential for piecing together the mechanism for how the TOC complex functions in protein import in apicomplexa and chloroplasts.

项目摘要 传染病每年在全世界引起广泛的疾病，是第二大疾病。 这是导致死亡的主要原因，特别是在不发达国家。随着多种药物的出现， 随着耐药菌株的出现，迫切需要新的、更有效的和更可持续的治疗方法。 包括在这些传染病中的是包括刚地弓形虫的顶复体， 恶性疟原虫，分别引起弓形虫病和疟疾的寄生虫。这些寄生虫 含有一种独特的类似质体的细胞器，称为顶质体，它含有四层膜，因此 已经进化出一个复杂的系统，用于通过这些膜输入和输出蛋白质。这些 重要的进出口机构是针对这些病原体的新型抗生素的理想目标。许多 在其他高等真核细胞器中， 叶绿体和线粒体，其中绝大多数基因是核编码的，因此必须 进口后，其中一种机制是外膜上保守的易位子 在拟南芥叶绿体复合体中，一个研究叶绿体生物学的模型系统。 TOC复合物主要由三种组分组成，Toc 33/34和Toc 159，两种都是GTP酶 含有将它们锚定到外膜中的N-末端跨膜螺旋，和Toc 75，一个16- 双股β-桶跨膜易位子。虽然已经提出了机械模型， TOC复杂的功能，他们仍然在很大程度上未经证实，由于缺乏结构 表征，这是需要缝合在一起的机械拼图的所有部分。在我们的研究中， 我们将使用生物物理学方法，X射线晶体学，冷冻电子显微镜和小角度X射线 分散到结构上和功能上表征这种专门的机械。我们的成果将填补 这是该领域长期存在的差距，对于拼凑TOC如何 顶复体和叶绿体中蛋白质输入的复杂功能。

项目成果

Divergence and Convergence: Complexity Emerges in Crystal Engineering from an 8-mer DNA.

• DOI: 10.1021/jacs.3c01941
• 发表时间: 2023-05
• 期刊: Journal of the American Chemical Society
• 影响因子: 15
• 作者: Jiemin Zhao;Cuizheng Zhang;Brandon Lu;R. Sha;N. Noinaj;C. Mao

The big BAM theory: An open and closed case?

BAM 大理论：开放式和封闭式案例？

• DOI: 10.1016/j.bbamem.2019.183062
• 发表时间: 2020
• 期刊: Biochimica et biophysica acta. Biomembranes
• 作者: Wu,Runrun;Stephenson,Robert;Gichaba,Abigail;Noinaj,Nicholas
• 通讯作者: Noinaj,Nicholas

Engineering DNA Crystals toward Studying DNA-Guest Molecule Interactions.

• DOI: 10.1021/jacs.3c00081
• 发表时间: 2023-02
• 期刊: Journal of the American Chemical Society
• 影响因子: 15
• 作者: Cuizheng Zhang;Jiemin Zhao;Brandon Lu;N. Seeman;R. Sha;N. Noinaj;C. Mao

Targeting BAM for Novel Therapeutics against Pathogenic Gram-Negative Bacteria.

针对病原革兰氏阴性细菌的新型治疗剂的靶向BAM。

• DOI: 10.3390/antibiotics12040679
• 发表时间: 2023-03-30
• 期刊: ANTIBIOTICS-BASEL
• 影响因子: 4.8
• 作者: Cottom, Claire Overly;Stephenson, Robert;Wilson, Lindsey;Noinaj, Nicholas
• 通讯作者: Noinaj, Nicholas

Plasticity within the barrel domain of BamA mediates a hybrid-barrel mechanism by BAM.

• DOI: 10.1038/s41467-021-27449-4
• 发表时间: 2021-12-08
• 期刊: Nature communications
• 影响因子: 16.6
• 作者: Wu R;Bakelar JW;Lundquist K;Zhang Z;Kuo KM;Ryoo D;Pang YT;Sun C;White T;Klose T;Jiang W;Gumbart JC;Noinaj N
• 通讯作者: Noinaj N