Protein Localization, Identification and Folding Core
Asma Nusrat, M.D.
Yatrik Shah, Ph.D.
John A. Williams, M.D., Ph.D.
Core Associate Director- Imaging
Alexey Nesvizhskii , Ph.D.
Core Associate Director- Proteomics
734-615-5510 or 734-647-3516
- Imaging Core Program (Service Request Form)
- Proteomics Core Program (Service Request Form)
- Protein Folding Core Program (Service Request Form)
- Consultation (Service Request Form)
1. Imaging Core Program
The imaging component, directed by Drs. Williams and Nusrat, utilizes a centralized facility supported by the University of Michigan for confocal and electron microscopy (EM) imaging. Instruction in the operation of confocal and electron microscopes will be provided by the Microscopy & Image Analysis Lab (MIL) facilities in the Medical School and the Morphology and Image Analysis Center (MIAC) facilities of the Michigan Diabetes Research and Training Center. Instruction as it applies to GI tissue will be provided by Drs. Nusrat and Williams, and by an experienced technician, Brad Nelson.
The Core will provide training in preparing frozen tissue samples, use of the PLIF Core cryostat to prepare frozen sections, immunohistochemistry, use of a fluorescence microscope and image processing. Instructions related to slide scanning will be provided by the ULAM Core director Robert Sigler. Aperio image analysis of scanned slides will be facilitated by Drs. Sigler at the ULAM facility (part of the service charge) and by Dr. Nusrat (who is an expert in the use of this imaging system). Core Service Request Form
2. Proteomics Core Program
The Proteomics program directed by Dr. Nesvizhskii and managed by Dr. Venky Basrur will utilize facilities located within the Medical School Department of Pathology on the fourth floor of Medical Science I building. This facility provides high-end mass spectrometry instrumentation dedicated for proteomic analysis. Presently PRF offers a variety of services including: 1) Simple protein identification from either gels or in-solution samples; 2) Interactome analysis with identification of protein-protein interaction(s) using immuno- or affinity-purified samples; 3) Shotgun proteomics. Mostly in-solution digestion followed by 1D (capable of resolving ~2000 proteins) or 2D LC-MS/MS (up to 6000-8000 proteins); 4) Post-translational modification analysis (e.g., phosphorylation, ubiquitylation, acetylation, methylation, glycosylation); 5) Differential proteome analysis using standard isotope labeling technology (SILAC, TMT etc.) or label-free approach; 6) Other: Non-routine applications of proteomic technology (e.g., unusual PTMs such as transamidation). Core Service Request Form
3. Protein Folding Core Program
The PFD services are facilitated by Dr. Omary, who serves as a member of the PFD Initiative Internal Advisory Committee. The key aspects of these services are its Resource Center that includes the PFD Reagent Library (which provides access, at no charge, to 85 different antibody samples for preliminary experiments) and 25 mouse genetic strains. The antibodies are directed to a range of ER stress and degradation pathways proteins, chaperones, apoptosis related proteins, and organelle markers. Because of the significant savings of needing to purchase antibodies for pilot experiments (typically $200-350 per antibody), this Core will provide our Center investigators at no cost a 20µL (typically 20µg) aliquot to use for pilot experiments. The mouse genetic model library include knockouts of secretory pathway genes such as SEC23A and B; SEC24A, B, C and D; SAR1A and B; and the Akita mouse. Core Service Request Form
As Co-directors of the PLIF Core Drs. Nusrat and Omary will meet monthly with the Core Management Committee consisting of the Associate Directors (Nesvizhskii, Williams) to discuss technologic updates and advances, member usage, priority access and changes in member subsidies to assess usage trends and member needs. Additionally, Dr. Nesvizhskii and Williams will meet weekly with the Research Scientist/Manager (Basrur) and Core Technician (Nelson) respectively to discuss Core experiments, potential problems or special requests, and turn-around times. The Core Directors, Associate Directors and personnel involved in Core operation will be available to consult with Center members on their research projects and Core use. After verifying that the research falls under the Center mission, the Co-directors will follow up with Core users to recommend interactions with the Program directors/associate directors and managers on an as-needed basis. This will be required for first-time users, regardless of whether they are early or established investigators. Experienced Core users can directly move forward with their service requests after proposal submission and brief review for Center relevance. If an applicant is denied Core services, the Directors will work with the applicant member to revise the application, e.g., more relevant approach, referral to other UMCGR Cores, or recommend additional experiments to be performed before Core usage. In summary, the Core Directors will assist member applicants with efficient GI-centric use of the PLIF Core resources. Core Service Request Form