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Clonable Nanoparticles

可克隆纳米颗粒

基本信息

批准号：
10378522
负责人：
Christopher Jeffries Ackerson
金额：
$ 29.46万
依托单位：
COLORADO STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-04-01 至 2024-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10378522
关键词：
Address Animals Area Artificial nanoparticles Binding Biological Biological Models Biological Process Biological Testing Biology Caliber Cell division Cells Centrifugation Chemicals Collaborations Color Colorado Complement Contrast Media Coupled Crystallization Cultured Cells DNA DNA Shuffling Data Development Directed Molecular Evolution Disease Disulfides Drug Metabolic Detoxication Dyes Electron Microscopy Electrons Elements Engineering Environment Enzymes Family Fatty acid glycerol esters Filament Fluorescence Green Fluorescent Proteins Growth Homologous Gene Image Imaging technology In Situ In Vitro Investigation Ions Knowledge Libraries Light Light Microscope Lighting Macromolecular Complexes Magnetic Resonance Imaging Magnetism Maintenance Mercury (II) reductase Metals Methods Minerals Modality Modeling Morphology Muscle Mutagenesis Optics Organism Oxidoreductase Pathologic Processes Peptide Library Peptides Phase Plasmids Property Proteins Public Health Radiation Accidents Randomized Research Roentgen Rays Sampling Selenite Selenium Shapes Site Skin Stains Synaptic Transmission System Technology Testing Tissues Variant Viral Viral Pathogenesis Whole Organism Work X-Ray Medical Imaging base bioimaging bone cell fixation enzyme activity experimental study imaging modality improved in vivo interest metagenomic sequencing microscopic imaging mutant nanoparticle nanoparticulate novel optical imaging particle physical property portability pyridine nucleotide receptor reconstitution small molecule tool

项目摘要

PROJECT SUMMARY The objective of this proposal is to address the contrast problem in images formed by X-Ray, electron, or other scattering based illumination modalities. Briefly, images are made by contrast. In other words, we only see (or acquire information) on things that are distinguishable from their background. In all forms of biological imaging, many things are visible, yet many other things remain camouflaged or indistinguishable from the background. For instance, in an X-ray, it's easy to see bones, but not so easy to see muscles, fat or skin. This is also true in microscopic images, where it's often easy to see the edges of cells, but much harder to see the details inside cells. Green Fluorescent Protein and related fluorescent proteins complement small molecule stains and dyes to essentially solve the contrast problem in optical imaging. For imaging based on X-rays or electrons, however, there are no clonable contrast agents. Clonable contrast (a GFP homolog) in X-Ray or electron-based imaging could be understood as a `clonable nanoparticle.' Such a nanoparticle would scatter incident radiation and would be made by a protein that can be genetically fused to other proteins of interest. Three discrete chemical activities are needed for such a clonable nanoparticle: (1) conversion of bioavailable inorganic ions to insoluble nanoparticulate form; (2) maintenance of the nanoparticle at the protein that synthesizes it; (3) size control of the nanoparticle, where 5nm diameter is suggested as an ideal size. We recently isolated a metalloid reductase that appears to partially fulfill each of these chemical requirements. We propose to build on this finding to create a pipeline that will produce many clonable nanoparticles of distinct size, shape or elemental composition. The most broadly useful clonable nanoparticles will simultaneously incorporate X-ray/electron scattering, magnetism and fluorescence. Such nanoparticles could serve as `universal clonable contrast agents' functioning in optical, MRI, X-Ray and electron imaging. Such a tool would greatly facilitate integrative and/or correlative multiscale bioimaging, integrating information from multiple imaging modalities. The proposed work will proceed in three specific aims. Candidate clonable nanoparticles will be identified and refined in Aims 1a and 1b. Refinement of our existing clonable nanoparticle candidate will proceed through directed evolution methods of saturation mutagenesis, DNA shuffling, and random mutagenesis. Isolation of additional variants and novel enzymes will begin with field samples collected in areas with persistent environmental metal contamination. Candidate clonable nanoparticles will be assessed in in vitro, in situ and in vivo within Aim 2 using 3 model and experimental systems we have identified to assess portability across species and suitability in in vivo, in situ, and in vitro chemical environments.

项目总结