Cysteine अवशेषों की सॉल्वेंट अभिगम्यता पर का विश्लेषण<em> मक्का rayado FINO वायरस</em> वायरस की तरह कण में उत्पादित<em> निकोटियाना benthamiana</em> पौधों और VLPs पेप्टाइड्स के पार से जोड़ने
Summary
सिस्टीन अवशेषों के की thiol समूह के विलायक पहुंच का विश्लेषण करने के लिए एक तरीका<em> मक्का rayado FINO वायरस</em> (MRFV) वायरस की तरह (VLPs) कणों पार से जोड़ने की एक पेप्टाइड प्रतिक्रिया द्वारा बाद में वर्णित है. विधि VLPs है कि विशिष्ट प्रतिक्रियाओं के लिए लक्ष्य हो सकता है की सतह पर कई रासायनिक समूहों की उपलब्धता का लाभ लेता है.
Abstract
Mimicking and exploiting virus properties and physicochemical and physical characteristics holds promise to provide solutions to some of the world’s most pressing challenges. The sheer range and types of viruses coupled with their intriguing properties potentially give endless opportunities for applications in virus-based technologies. Viruses have the ability to self- assemble into particles with discrete shape and size, specificity of symmetry, polyvalence, and stable properties under a wide range of temperature and pH conditions. Not surprisingly, with such a remarkable range of properties, viruses are proposed for use in biomaterials 9, vaccines 14, 15, electronic materials, chemical tools, and molecular electronic containers4, 5, 10, 11, 16, 18, 12.
In order to utilize viruses in nanotechnology, they must be modified from their natural forms to impart new functions. This challenging process can be performed through several mechanisms including genetic modification of the viral genome and chemically attaching foreign or desired molecules to the virus particle reactive groups 8. The ability to modify a virus primarily depends upon the physiochemical and physical properties of the virus. In addition, the genetic or physiochemical modifications need to be performed without adversely affecting the virus native structure and virus function. Maize rayado fino virus (MRFV) coat proteins self-assemble in Escherichia coli producing stable and empty VLPs that are stabilized by protein-protein interactions and that can be used in virus-based technologies applications 8. VLPs produced in tobacco plants were examined as a scaffold on which a variety of peptides can be covalently displayed 13. Here, we describe the steps to 1) determine which of the solvent-accessible cysteines in a virus capsid are available for modification, and 2) bioconjugate peptides to the modified capsids. By using native or mutationally-inserted amino acid residues and standard coupling technologies, a wide variety of materials have been displayed on the surface of plant viruses such as, Brome mosaic virus 3, Carnation mottle virus 12, Cowpea chlorotic mottle virus 6, Tobacco mosaic virus 17, Turnip yellow mosaic virus 1, and MRFV 13.
Protocol
Representative Results
Discussion
यहाँ प्रस्तुत विधि प्रतिक्रियाशील cysteines संयंत्र उत्पादन VLPs की सतह पर के रूप में के रूप में अच्छी तरह से अन्य प्रोटीन परिसरों पर वर्तमान के एक बहुत ही संवेदनशील और तेजी से विश्लेषण के लिए सक्षम बनाता है. Male…
Disclosures
The authors have nothing to disclose.
Materials
| Name of the reagent | Company | Catalog number | Comments |
| Thinwall, Ultra-Clear Tubes | Beckman | 344059 | |
| mMESSAGE mMACHINE T7 Kit | Life Tecnologies | AM1344M | |
| Fluorescein-5-Maleimide | Thermo Scientific Life Technologies | 46130 F150 | 46130 is out of order substitute with F150 |
| Pierce Biotin Quantitation Kit | Thermo Scientific | 28005 | |
| EZ-Link Maleimide-PEG2-Biotin, No-Weigh Format | Thermo Scientific | 21901 | |
| SM(PEG)n Crosslinkers | Thermo Scientific | 22107 | |
| 10-20% Tris-Glycine gel | Invitrogen | EC61352 | |
| Laemmli Buffer | Bio-Rad | 1610737 | |
| Tris Glycine SDS Running Buffer | Invitrogen | LC2675 | |
| Tris Glycine Transfer Buffer | Invitrogen | LC3675 | |
| Nitrocellulose Membrane Filter Paper Sandwich | Invitrogen | LC2001 | |
| Phosphatase Labeled Affinity Purified Antibody to Rabbit IgG | Kirkegaard and Perry Laboratories | 0751516 | |
| NBT/BCIP Phosphatase Substrate | Kirkegaard and Perry Laboratories | 508107 |
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