scholarly journals Fluorescent Nanodiamonds Encapsulated by Cowpea Chlorotic Mottle Virus (CCMV) Proteins for Intracellular 3D-Trajectory Analysis

2021 ◽  
Author(s):  
Yingke Wu ◽  
Shuqin Cao ◽  
Md Noor A Alam ◽  
Marco Raabe ◽  
Sandra Michel-Souzy ◽  
...  
Keyword(s):  
Trajectory Analysis ◽  
Living Cells ◽  
Transport Processes ◽  
Mottle Virus ◽  
Cowpea Chlorotic Mottle Virus ◽  
Fundamental Parameters ◽  
Chlorotic Mottle Virus ◽  
Chlorotic Mottle ◽  
Fluorescent Nanodiamonds

Long-term tracking of nanoparticles to resolve intracellular structures and motions is essential to elucidate fundamental parameters as well as transport processes within living cells. Fluorescent nanodiamond (ND) emitters provide cell...

scholarly journals Fluorescent Nanodiamonds Encapsulated by Cowpea Chlorotic Mottle Virus (CCMV) Proteins for Intracellular 3D-Trajectory Analysis

2021 ◽  
Author(s):  
Yingke Wu ◽  
Shuqin Cao ◽  
Md Noor A Alam ◽  
Marco Raabe ◽  
Sandra Michel-Souzy ◽  
...  
Keyword(s):  
Cell Biology ◽  
Colloidal Stability ◽  
Living Cells ◽  
Transport Processes ◽  
Mottle Virus ◽  
Cowpea Chlorotic Mottle Virus ◽  
Physiological Conditions ◽  
Chlorotic Mottle Virus ◽  
Chlorotic Mottle

Long-term tracking of nanoparticles to resolve intracellular structures and motions is essential to elucidate fundamental parameters as well as transport processes within living cells. Fluorescent nanodiamond (ND) emitters provide cell compatibility and very high photostability. However, high stability, biocompatibility, and cellular uptake of these fluorescent NDs under physiological conditions are required for intracellular applications. Herein, highly stable NDs encapsulated with Cowpea chlorotic mottle virus capsid proteins (ND-CP) are prepared. A thin capsid protein layer is obtained around the NDs, which imparts reactive groups and high colloidal stability, while retaining the opto-magnetic properties of the coated NDs as well as the secondary structure of CPs adsorbed on the surface of NDs. In addition, the ND-CP shows excellent biocompatibility both in vitro and in vivo. Long-term 3D trajectories of the ND-CP with fine spatiotemporal resolutions are recorded; their intracellular motions are analyzed by different models, and the diffusion coefficient are calculated. The ND-CP with its brilliant optical properties and stability under physiological conditions provides us with a new tool to advance the understanding of cell biology, e.g., endocytosis, exocytosis, and active transport processes in living cells as well as intracellular dynamic parameters. <br>

scholarly journals Fluorescent Nanodiamonds Encapsulated by Cowpea Chlorotic Mottle Virus (CCMV) Proteins for Intracellular 3D-Trajectory Analysis

2021 ◽  
Author(s):  
Yingke Wu ◽  
Shuqin Cao ◽  
Md Noor A Alam ◽  
Marco Raabe ◽  
Sandra Michel-Souzy ◽  
...  
Keyword(s):  
Cell Biology ◽  
Colloidal Stability ◽  
Living Cells ◽  
Transport Processes ◽  
Mottle Virus ◽  
Cowpea Chlorotic Mottle Virus ◽  
Physiological Conditions ◽  
Chlorotic Mottle Virus ◽  
Chlorotic Mottle

Long-term tracking of nanoparticles to resolve intracellular structures and motions is essential to elucidate fundamental parameters as well as transport processes within living cells. Fluorescent nanodiamond (ND) emitters provide cell compatibility and very high photostability. However, high stability, biocompatibility, and cellular uptake of these fluorescent NDs under physiological conditions are required for intracellular applications. Herein, highly stable NDs encapsulated with Cowpea chlorotic mottle virus capsid proteins (ND-CP) are prepared. A thin capsid protein layer is obtained around the NDs, which imparts reactive groups and high colloidal stability, while retaining the opto-magnetic properties of the coated NDs as well as the secondary structure of CPs adsorbed on the surface of NDs. In addition, the ND-CP shows excellent biocompatibility both in vitro and in vivo. Long-term 3D trajectories of the ND-CP with fine spatiotemporal resolutions are recorded; their intracellular motions are analyzed by different models, and the diffusion coefficient are calculated. The ND-CP with its brilliant optical properties and stability under physiological conditions provides us with a new tool to advance the understanding of cell biology, e.g., endocytosis, exocytosis, and active transport processes in living cells as well as intracellular dynamic parameters. <br>

Expression and Self Assembly of Cowpea Chlorotic Mottle Virus Capsid Proteins inPichia pastorisand Encapsulation of Fluorescent Myoglobin

2011 ◽  
Vol 1317 ◽  
Author(s):  
Yuanzheng Wu ◽  
Hetong Yang ◽  
Hyun-Jae Shin
Keyword(s):  
Coat Protein ◽  
Large Scale ◽  
Materials Science ◽  
Cesium Chloride ◽  
Size Exclusion ◽  
Mottle Virus ◽  
Cowpea Chlorotic Mottle Virus ◽  
Chlorotic Mottle Virus ◽  
Chlorotic Mottle

Abstract:Cowpea chlorotic mottle virus (CCMV) has been a model system for virus studies for over 40 years and now is considered to be a perfect candidate as nanoplatform for applications in materials science and medicine. The ability of CCMV to self assemblein vitrointo virus-like particles (VLPs) or capsids makes an ideal reaction vessel for nanomaterial synthesis and entrapment. Here we report expression of codon optimized CCMV coat protein inPichia pastorisand production of self assembled CCMV VLPs by large-scale fermentation. CCMV coat protein gene (573 bp) was synthesized according to codon preference ofP. pastorisand cloned into pPICZA vector. The recombinant plasmid pPICZA-CP was transformed intoP. pastorisGS115 by electroporation. The resulting yeast colonies were screened by PCR and analyzed for protein expression by SDS-PAGE. After large-scale fermentation CCMV coat protein yields reached 4.8 g L−1. The CCMV VLPs were purified by modified PEG precipitation followed by cesium chloride density gradient ultracentrifugation, and then analyzed by size exclusion fast performance liquid chromatography (FPLC), UV spectrometry and transmission electron microscopy. Myoglobin was used as a model protein to be encapsulated in CCMV VLPs. The fluorescence spectroscopy showed that inclusion of myoglobin had occurred. The results indicated the production of CCMV capsids byP. pastorisfermentation now available for utilization in pharmacology or nanotechnology fields.

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