This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Severe acute respiratory syndrome coronavirus (SARS CoV) is an important emerging disease-causing agent, but very little is known about how SARS CoV causes disease. We propose a model in which the SARS CoV nucleocapsid (N) protein is proposed to localize to the nucleolus of the host cell during virus infection and alter nucleolar function: an important viral strategy designed to harm the host cell while benefiting the replicating virus. The analysis of the SARS virus N protein shows several regions that are predicted to be involved in the localization of the N protein to the nucleolus. The first part of this project is to determine if SARS CoV N protein does in fact localize to the nucleolus, and to characterize the protein domains involved in transporting N into the nucleus and nucleolus. The second part, in collaboration with the COBRE Core C Protein Purification Group, is to construct a detailed structural model of the N protein, which will be used in future studies to better understand the mechanism of how the SARS virus causes disease and identify targets for potential antiviral drugs. To study the N protein in SARS CoV-infected cells, we developed several monoclonal antibody (mAb) reagents. To our surprise, confocal microscopy of infected cells stained with the N-specific mAb, 46-4, showed clearly that N protein fails to localize in the nucleus or nucleolus as originally predicted. Similar results were obtained for N expressed alone or tagged with enhanced green fluorescent protein (EGFP). Lack of nuclear transport activity was an unexpected finding, but was confirmed in several experiments. Even though the N protein of SARS CoV is predicted to be a nuclear protein, it does not localize to the nucleolus and is primarily retained in the cytoplasm. We are currently investigating the structure of N as a means to determine why it fails to localize to the nucleus. An important side benefit of this work is the creation of several antibody reagents, which will further enhance the understanding of SARS CoV N protein structure and function and prove useful for the development of N protein-specific tests for the diagnosis of SARS CoV infection.
| Garabedian, Alyssa; Baird, Matthew A; Porter, Jacob et al. (2018) Linear and Differential Ion Mobility Separations of Middle-Down Proteoforms. Anal Chem 90:2918-2925 |
| Jeanne Dit Fouque, Kevin; Garabedian, Alyssa; Porter, Jacob et al. (2017) Fast and Effective Ion Mobility-Mass Spectrometry Separation of d-Amino-Acid-Containing Peptides. Anal Chem 89:11787-11794 |
| Alaofi, Ahmed; Farokhi, Elinaz; Prasasty, Vivitri D et al. (2017) Probing the interaction between cHAVc3 peptide and the EC1 domain of E-cadherin using NMR and molecular dynamics simulations. J Biomol Struct Dyn 35:92-104 |
| Pang, Xiao-Yan; Wang, Suya; Jurczak, Michael J et al. (2017) Retinol saturase modulates lipid metabolism and the production of reactive oxygen species. Arch Biochem Biophys 633:93-102 |
| McNiff, Michaela L; Chadwick, Jennifer S (2017) Metal-bound claMP Tag inhibits proteolytic cleavage. Protein Eng Des Sel 30:467-475 |
| Budiardjo, S Jimmy; Licknack, Timothy J; Cory, Michael B et al. (2016) Full and Partial Agonism of a Designed Enzyme Switch. ACS Synth Biol 5:1475-1484 |
| O'Neil, Pierce; Lovell, Scott; Mehzabeen, Nurjahan et al. (2016) Crystal structure of histone-like protein from Streptococcus mutans refined to 1.9?Å resolution. Acta Crystallogr F Struct Biol Commun 72:257-62 |
| Gowthaman, Ragul; Miller, Sven A; Rogers, Steven et al. (2016) DARC: Mapping Surface Topography by Ray-Casting for Effective Virtual Screening at Protein Interaction Sites. J Med Chem 59:4152-70 |
| Kumar, Ritesh; Qi, Yifei; Matsumura, Hirotoshi et al. (2016) Replacing Arginine 33 for Alanine in the Hemophore HasA from Pseudomonas aeruginosa Causes Closure of the H32 Loop in the Apo-Protein. Biochemistry 55:2622-31 |
| Meekins, David A; Zhang, Xin; Battaile, Kevin P et al. (2016) 1.45?Å resolution structure of SRPN18 from the malaria vector Anopheles gambiae. Acta Crystallogr F Struct Biol Commun 72:853-862 |
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