Enabling Technologies and Tools
The primary rationale for the PCHPI organization is to maximize interdisciplinary collaboration and access to the PCHPI resources, while maintaining focused expertise in technical disciplines. The Center is organized around four methodological cores (Computational, Cryo-EM/ET, NMR, and HIV Virology) and three scientific projects (1. Maturation; 2. Trafficking, Restriction and Nuclear Entry; and 3. Integration). The cores are complemented by the expertise of our collaborators in several important areas. Together, close collaboration among the research groups and cores allows the PCHPI to take a "hybrid" approach to answer important structural, biochemical, and biophysical questions about HIV-1.
Computational Core
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- Derivation of full-scale models that integrate experimental structural data
- Design and execution of large-scale and long-timescale simulations
- Design and execution of mechanistic studies based on free energy calculations and steered molecular dynamics
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Cryo-EM & Cryo-ET Core
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- Cryo-Electron Single Particle Analysis
- Cryo-Electron Sub-Tomogram Averaging
- Micro-Electron Diffraction (micro-ED) of nanocrystals and crystal lamellae
- Cryo-Focused Ion Beam/Scanning EM for cryo-ET of in-cell lamellae and micro-ED of crystal lamellae
- Cryo-FIB/SEM 3D volume imaging of whole frozen cell/tissue
- Correlative cryo-flourescence microscopy and cryo-ET (CLEM)
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NMR spectroscopy Core
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- Screening of recombinant proteins to evaluate their biophysical properties, structural integrity and suitability for further structural determinations
- Determination of structures and dynamics of protein assemblies and their complexes with small molecules by MAS NMR
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HIV Virology Core
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- HIV-1 infectivity assays
- Purification of HIV-1 cores
- Imaging of early events (Confocal and TIRF microscopy)
- Immunoblot analysis of HIV-1 particles
- TEM analysis of HIV-1 particles
- Quantitative analysis of reverse transcription, nuclear import, and integration in target cells
- HIV-1 integration site analysis
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