Infections with human immunodeficiency virus (HIV) and hepatitis C virus (HCV) are among the most significant causes of human morbidity and mortality. Worldwide, there are more than 40 million (HIV) and 170 million (HCV) people infected with these viruses. In the United States, 0.6% (HIV) and 1.7% (HCV) of the population is infected. More than 25% of those infected with HIV are co-infected with HCV (up to 90% of i.v. drug users). Despite the success of highly active anti-retroviral therapy, there is an urgent need to develop entirely new classes of anti-HIV drugs targeted to novel receptors in order to ensure different susceptibilities to the development of resistance. Current therapies for HCV are grossly inadequate, and multiple first-generation anti- HCV drugs are needed. Both HIV-1 and HCV have a viroporin, a small membrane protein with ion channel activity and a potential drug target. The proposed research builds upon recent results, and represents a substantial expansion of our structural studies of membrane proteins. Determining the structures of Vpu (Virus protein """"""""u"""""""") from HIV-1 and the p7 protein from HCV are essential in order to understand the molecular mechanisms of their biological activities and to nucleate the discovery of drugs that interfere with these activities. Our NMR methods are particularly well suited for determining the structures of these proteins in their native environment of phospholipid bilayers. And since our approach is unaffected by the molecular mass of the polypeptides, studies of isotopically labeled Vpu bound to unlabeled cellular partners are no more difficult than for Vpu alone. We will explore the application of SAR (structure activity relationships) by NMR to Vpu and p7 as examples of membrane-bound receptors and to identify potential binding sites for drugs.

Public Health Relevance

Infections with human immunodeficiency virus (HIV) and hepatitis C virus (HCV) are among the most significant causes of human morbidity and mortality. Worldwide, there are more than 40 million (HIV) and 170 million (HCV) people infected with these viruses. In the United States, 0.6% (HIV) and 1.7% (HCV) of the population is infected. More than 25% of those infected with HIV are co-infected with HCV (up to 90% of i.v. drug users). HIV is a lentivirus in the Retroviridae family. It causes the acquired immunodeficiency syndrome (AIDS) by infecting CD4+ T and other cells of the immune system; the resulting decline of immune functions means that opportunistic infections, such as tuberculosis and Kaposi's sarcoma, and co-infections, such as HCV, have devastating effects on human health. HAART (highly active antiretroviral therapy) is effective in many patients; however, the genetic variability of HIV means that drugs against additional molecular targets are needed to combat resistance that inevitably develops in the course of treating a disease in which the genome of the causative agent is integrated into the host DNA. The proposed studies will provide structural information about Vpu, a key protein in the viral lifecycle that has the potential to be a new target for the development of drugs to treat AIDS. HCV is a member of the Flaviviridae family of enveloped single-stranded RNA viruses that mainly infects hepatocyte. Most HCV infections are persistent and lead to liver diseases, including hepatitis, cirrhosis, and hepatocellular carcinoma. Consequently, HCV is the leading indicator for liver transplantation in the Nation. Current therapy for HCV infections consists of the administration of pegylated interferon and ribivirin and has many limitations; not only is it lengthy, expensive and poorly tolerated, but also it works in only a fraction of cases. The development of effective anti-HCV drugs is a high priority, especially because of the difficulties encountered in treating patients co-infected with HIV and HCV. The proposed studies will provide structural information about p7, a key protein in the viral lifecycle that has the potential to be a new target for the development of drugs to treat hepatitis C. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM066978-05A2
Application #
7508672
Study Section
Special Emphasis Panel (ZRG1-BCMB-B (02))
Program Officer
Wehrle, Janna P
Project Start
2003-07-01
Project End
2012-06-30
Budget Start
2008-07-07
Budget End
2009-06-30
Support Year
5
Fiscal Year
2008
Total Cost
$303,961
Indirect Cost
Name
University of California San Diego
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
804355790
City
La Jolla
State
CA
Country
United States
Zip Code
92093
Berkamp, Sabrina; Park, Sang Ho; De Angelis, Anna A et al. (2017) Structure of monomeric Interleukin-8 and its interactions with the N-terminal Binding Site-I of CXCR1 by solution NMR spectroscopy. J Biomol NMR 69:111-121
Opella, Stanley J; Marassi, Francesca M (2017) Applications of NMR to membrane proteins. Arch Biochem Biophys 628:92-101
Park, Sang Ho; Berkamp, Sabrina; Radoicic, Jasmina et al. (2017) Interaction of Monomeric Interleukin-8 with CXCR1 Mapped by Proton-Detected Fast MAS Solid-State NMR. Biophys J 113:2695-2705
Yao, Yong; Dutta, Samit Kumar; Park, Sang Ho et al. (2017) High resolution solid-state NMR spectroscopy of the Yersinia pestis outer membrane protein Ail in lipid membranes. J Biomol NMR 67:179-190
Das, Bibhuti B; Opella, Stanley J (2016) Simultaneous cross polarization to (13)C and (15)N with (1)H detection at 60kHz MAS solid-state NMR. J Magn Reson 262:20-26
Opella, Stanley J (2015) Solid-state NMR and membrane proteins. J Magn Reson 253:129-37
Das, Bibhuti B; Park, Sang Ho; Opella, Stanley J (2015) Membrane protein structure from rotational diffusion. Biochim Biophys Acta 1848:229-45
Opella, Stanley J (2015) Relating structure and function of viral membrane-spanning miniproteins. Curr Opin Virol 12:121-5
Lewinski, Mary K; Jafari, Moein; Zhang, Hua et al. (2015) Membrane Anchoring by a C-terminal Tryptophan Enables HIV-1 Vpu to Displace Bone Marrow Stromal Antigen 2 (BST2) from Sites of Viral Assembly. J Biol Chem 290:10919-33
Park, Sang Ho; Wang, Vivian S; Radoicic, Jasmina et al. (2015) Paramagnetic relaxation enhancement of membrane proteins by incorporation of the metal-chelating unnatural amino acid 2-amino-3-(8-hydroxyquinolin-3-yl)propanoic acid (HQA). J Biomol NMR 61:185-96

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