Clonable Nanoparticles

可克隆纳米颗粒

基本信息

批准号：
8502254
负责人：
Christopher Jeffries Ackerson
金额：
$ 20.67万
依托单位：
COLORADO STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-07-01 至 2015-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8502254
关键词：
Amino Acids Antibiotics Area Bacteria Biocompatible Materials Biological Biological Assay Biology Cell Extracts Cells Chemicals Chemistry Chimeric Proteins Codon Nucleotides Complement Complex Cryoelectron Microscopy Custom Development Diagnosis Disease Electron Microscope Electron Microscopy Electrons Environment Exploratory/Developmental Grant Ferritin Filament Freezing Funding Gene Expression Genotype Gold Green Fluorescent Proteins In Situ In Vitro Individual Ions Learning Length Libraries Light Microscope Link Literature Magnetic Resonance Imaging Magnetism Mediating Messenger RNA Metals Microscope Microscopy Microtubules Minor Modality Modification Molecular Nobel Prize Optics Organism Oxidation-Reduction Peptide Fragments Peptide Library Peptide Phage Display Library Peptides Phage Display Phenotype Photons Play Property Protein Fragment Proteins Public Health Reading Reporter Genes Resolution Ribosomes Role Sampling Shapes Site Solutions Staining method Stains Structure System Techniques Technology Testing Time Tissues Tomogram Transition Elements Tubulin Visual Work Yeasts analog base cell fixing directed evolution disilver oxide electron crystallography fluorescence microscope germanium oxide in vivo iron oxide light microscopy magnetite ferrosoferric oxide metal oxide nanoparticle novel particle phytochelatin protein aminoacid sequence research study small molecule success tomography

项目摘要

DESCRIPTION (provided by applicant): The objective of this exploratory (R21) proposal is to identify the most promising strategy for solving the contrast problem in electron microscopy. In all microscopes, whether illuminated by photons or electrons, biological tissues are essentially transparent. Green Fluorescent Protein (GFP) and related fluorescent proteins have complemented small molecule stains to essentially solve the contrast problem in optical microscopy. For electron microscopy, however, there are no 'clonable' contrast markers analogous to GFP. Herein we propose to explore several peptide or small protein based strategies for nucleating, catalyzing or mediating the formation of electron dense inorganic nanoparticles which may allow the localization of individual proteins in macromolecular and whole cell electron microscopy. The proposed work is significant for four areas of inquiry. First and most significantly, a robust clonable nanoparticle will revolutionize electron microscopy, making it significant for all forms of biological EM. Second, we will study FtsZ, the prokaryotic tubulin analog as an initial target. Information we learn about the intrcellular structure and distribution of FtsZ is significant for the development of antibiotics that target FtsZ. Third, we will, in finding novel peptides or small proteins that interact with metal ions or coordination complexes learn more about the important and sometimes poorly understood interactions of metals and biomolecules. Fourth just as GFP can act as a 'reporter gene' for reading optically for gene expression in tissues and organisms, a genetically encodable magnetic nanoparticle may also serve as a widely used 'reporter gene' for detecting gene expression by MRI or CT. The proposed work will proceed in three specific aims. All candidate 'clonable nanoparticle' peptides or proteins identified in these specific aims will be evaluated in vitro, in situ and in vvo in an FtsZ based structural electron microscopy assay. Aims one and two use the directed evolution techniques of phage display and ribosome display, respectively, to isolate peptide sequences capable of provoking the formation of magnetic iron oxide nanoparticles. Aim 2 is technically more challenging but is more likely to yield a universally useful clonable nanoparticle than Aim 1, which may yield clonable nanoparticles that function in situ and in vitro, but not in vivo. Aim 3 is to assay naturally occurring peptides, proteins and relevant protein fragments, as well as peptides isolated by others, for comparison to the peptides we isolate in aims 1 and 2. The proposal identifies why existing 'known' peptides and proteins fail to function as EM contrast markers, and suggests modifications that we may make to these known peptides and proteins to adapt them as EM contrast markers. The public health significance of the development of the proposed enabling technologies will derive from both more comprehensive structural information on normal and diseased cells, and also from greater understanding of FtsZ biology, which may enable development of new antibiotics.

描述（由申请人提供）：该探索性（R21）提案的目的是确定解决电子显微镜对比度问题的最有前途的策略。在所有显微镜中，无论是通过光子还是电子照明，生物组织基本上都是透明的。绿色荧光蛋白（GFP）及相关荧光蛋白与小分子染色剂相辅相成，从根本上解决了光学显微镜中的对比度问题。然而，对于电子显微镜，没有类似于 GFP 的“可克隆”对比标记。在此，我们建议探索几种基于肽或小蛋白质的策略，用于成核、催化或介导电子致密无机纳米粒子的形成，这可能允许在大分子和全细胞电子显微镜中定位单个蛋白质。拟议的工作对于四个研究领域具有重要意义。首先也是最重要的是，强大的可克隆纳米颗粒将彻底改变电子显微镜，使其对所有形式的生物电子显微镜具有重要意义。其次，我们将研究 FtsZ（原核微管蛋白类似物）作为初始靶点。我们了解的有关 FtsZ 细胞内结构和分布的信息对于开发针对 FtsZ 的抗生素具有重要意义。第三，通过寻找与金属离子或配位络合物相互作用的新型肽或小蛋白质，我们将更多地了解金属和生物分子之间重要的、有时人们知之甚少的相互作用。第四，正如 GFP 可以充当“报告基因”，用于光学读取组织和生物体中的基因表达一样，基因可编码的磁性纳米颗粒也可以充当广泛使用的“报告基因”，用于通过 MRI 或 CT 检测基因表达。拟议的工作将朝着三个具体目标进行。在这些特定目标中鉴定的所有候选“可克隆纳米颗粒”肽或蛋白质都将在基于 FtsZ 的结构电子显微镜测定中进行体外、原位和体内评估。目标一和二分别使用噬菌体展示和核糖体展示的定向进化技术来分离能够引发磁性氧化铁纳米粒子形成的肽序列。目标 2 在技术上更具挑战性，但更有可能产生普遍有用的可克隆纳米颗粒比目标 1 更好，它可能产生可在原位和体外发挥作用但在体内不起作用的可克隆纳米颗粒。目标 3 是分析天然存在的肽、蛋白质和相关蛋白质片段，以及其他人分离的肽，与我们在目标 1 和 2 中分离的肽进行比较。该提案确定了现有“已知”肽和蛋白质无法发挥作用的原因作为 EM 对比标记物，并建议我们可以对这些已知的肽和蛋白质进行修改，以使其适合作为 EM 对比标记物。所提议的使能技术的开发对公共卫生的意义将来自于正常细胞和患病细胞的更全面的结构信息，以及对 FtsZ 生物学的更深入的了解，这可能有助于开发新的抗生素。

项目成果

期刊论文数量（5）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（2）

Sensitive, selective analysis of selenium oxoanions using microchip electrophoresis with contact conductivity detection.

DOI：
10.1021/ac502013k
发表时间：
2014-08-19
期刊：
ANALYTICAL CHEMISTRY
影响因子：
7.4
作者：
Noblitt, Scott D.;Staicu, Lucian C.;Ackerson, Christopher J.;Henry, Charles S.
通讯作者：
Henry, Charles S.

Radicals Are Required for Thiol Etching of Gold Particles.

DOI：
10.1002/anie.201502934
发表时间：
2015-08-03
期刊：
Angewandte Chemie (International ed. in English)
影响因子：
0
作者：
Dreier TA;Ackerson CJ
通讯作者：
Ackerson CJ

Conformation and dynamics of the ligand shell of a water-soluble Au102 nanoparticle.

DOI：
10.1038/ncomms10401
发表时间：
2016-01-21
期刊：
Nature communications
影响因子：
16.6
作者：
Salorinne K;Malola S;Wong OA;Rithner CD;Chen X;Ackerson CJ;Häkkinen H
通讯作者：
Häkkinen H

Radiofrequency heating pathways for gold nanoparticles.

DOI：
10.1039/c4nr00464g
发表时间：
2014-08-07
期刊：
Nanoscale
影响因子：
6.7
作者：
Collins CB;McCoy RS;Ackerson BJ;Collins GJ;Ackerson CJ
通讯作者：
Ackerson CJ

Polymorphism in magic-sized Au144(SR)60 clusters.

DOI：
10.1038/ncomms11859
发表时间：
2016-06-14
期刊：
Nature communications
影响因子：
16.6
作者：
Jensen KM;Juhas P;Tofanelli MA;Heinecke CL;Vaughan G;Ackerson CJ;Billinge SJ
通讯作者：
Billinge SJ