Ryan N. Willhite Week 7: Difference between revisions
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** Automatic concatenation of unmodelled density using PRISM | ** Automatic concatenation of unmodelled density using PRISM | ||
* Measure deviation | * Measure deviation | ||
[[Media:JOURNALCLUB.ppt|Powerpoint Presentation]] | |||
==Unfamiliar Words: == | ==Unfamiliar Words: == | ||
#glycoprotein | #glycoprotein-are proteins that contain oligosaccharide chains covalently bound to polypeptide side chains. | ||
#chemokine | #chemokine-One of a large group of proteins that act as lures and were first found attracting white blood cells. | ||
#cavity-laden | #cavity-laden- | ||
#oligomeric | #oligomeric-A polymer that consists of two, three, or four monomers. | ||
#virion | #virion-A complete viral particle, consisting of RNA or DNA surrounded by a protein shell and constituting the infective form of a virus. | ||
#fusogenic | #fusogenic-cause cell fusion | ||
#interfacial cavities | #interfacial cavities | ||
#haemagglutinin | #haemagglutinin-involved in influenza. clogs red blood cells together. | ||
#steric occlusion | #steric occlusion- as a General Pro Region Inhibition Mechanism | ||
#cryoprotectant | *pubs.acs.org/cgi-bin/jtext?bichaw/36/i13/abs/bi962341o | ||
#cryoprotectant- protects biological tissue from freezing. | |||
*[[BIOL398-01/S10:Week 7|Week 7 Assignment Page]] | *[[BIOL398-01/S10:Week 7|Week 7 Assignment Page]] |
Latest revision as of 00:53, 8 March 2010
Outline
I. Introduction
- Entry of HIV involves a sequential interaction of
- the envelope glycoprotein (gp120)
- CD4 glycoprotein
- chemokine receptor (primary receptor)
- CD4i (antibodies that block gp120-CD4 complexes to the chemokine receptor)
- CCR5 and CXCR4 for HIV-1 (secondary receptors)
- CD4 binding induces conformational changes in the gp120
- Entry of HIV is mediated by envelope glycoproteins
- 5 variable regions
- Variable and non variable regions are glycosylated
- V3 loop determines specificity
- Gp41 (transmembrane coat proteins) variants found in all enveloped viruses
- N-terminal fusion peptides which participate in membrane fusion
- Enveloped viruses tend to be characteristic in entry
- Direct membrane penetration (HIV)
- HIV causes destruction of CD4 T cells which ultimately leads to AIDS.
II. Purpose of this Study
- Gp120 glycoprotein has important role in
- receptor binding
- interactions with neutralizing antibodies
- Information about the gp120 structure is important for understanding HIV infection
- Assist in designing therapeutic strategies.
- Overall purpose is to observe the mechanism of HIV entry and intervene
III. Determining the Structure
- Devised a crystallization strategy that modified the protein surface
- Obtained crystals of
- a ternary complex composed of a truncated form of gp120
- the N-terminal two domains (DID2) of CD4
- Fab from the human neutralizing monoclonal antibody 17
- The ternary structure was solved by
- combinations of molecular replacement
- isomorphous replacement
- density modification techniques
IV. Methods
- Protein Production, crystallization and data collection
Diffraction data were collected at beamline X4A using phosphorimaging plates and Fuji BAS2000 scanner
- Structure determination
- To locate Fab 17b in the ternary complex crystals, rotational searches with 52 different Fab models were made with program MERLOT*
- Crystals soaked in over 20 different heavy atom compounds
- Isomorphous replacement phasing*
- Automatic concatenation of unmodelled density using PRISM
- Measure deviation
Unfamiliar Words:
- glycoprotein-are proteins that contain oligosaccharide chains covalently bound to polypeptide side chains.
- chemokine-One of a large group of proteins that act as lures and were first found attracting white blood cells.
- cavity-laden-
- oligomeric-A polymer that consists of two, three, or four monomers.
- virion-A complete viral particle, consisting of RNA or DNA surrounded by a protein shell and constituting the infective form of a virus.
- fusogenic-cause cell fusion
- interfacial cavities
- haemagglutinin-involved in influenza. clogs red blood cells together.
- steric occlusion- as a General Pro Region Inhibition Mechanism
- pubs.acs.org/cgi-bin/jtext?bichaw/36/i13/abs/bi962341o
- cryoprotectant- protects biological tissue from freezing.