Multi-Drug Resistant HIV-1 Protease
The current treatment used to control HIV-1 replication in affected patients in the United States is combination therapy, otherwise known as Highly Active Antiretroviral Therapy (HAART). The use of at least three antiretroviral medications at the same time, either two nucleoside inhibitors plus a protease inhibitor, or two nucleoside inhibitors plus a non-nucleoside inhibitor, is recommended by the National Institutes of Health . As a consequence of HIV-1 mutations, drug resistance, a major factor in reduced drug efficacy, is an issue. Past research analysis by our laboratory, using structural modeling and X-ray crystallography, shows that the multi-drug resistant (MDR) HIV-1 protease has an expanded active site cavity, shorter amino acids side chains protruding into the active site that bind to substrate, wider flaps on the perimeter of the protease to keep the active site open in a wider format, and completely different protease-ligand interactions, resulting in a change in binding affinity. Thus, MDR HIV-1 is a new antiviral target. The determination of the MDR HIV-1 protease substrate envelope is essential to determining potent substrate inhibitors at its active site. Our current project builds upon our previous findings and focuses on the synthesis of reduced peptides, exploring their interaction and affinity to the expanded active site, and the design of drugs that would be more affective in a multi-drug resistant HIV-1 protease.
Hepatitis C NS3/4A Protease
The hepatitis C virus (HCV) causes a chronic liver disease in affected individuals around the world. Populations in different geographical locations will contain variants of HCV, known as viral “genotypes.” The NS3 gene of the HCV contains the coding information for a NS3 bifunctional protease/helicase. Cofactor NS4A is required for the active NS3 conformational state. The NS3 protease cleaves other HCV non-structural proteins (including its own cofactor) aiding in the succession of the virus. In conjunction with Vertex Pharmaceuticals, this project targets the inhibition efficacy of a pharmaceutical drug that is currently directed at the NS3 protease in HCV genotype 1. We will test its inhibition efficacy in HCV genotypes 4,5, and 6. We will also perform structure-functions studies on all NS3 apoprotease variants well as protease-ligand co-crystallization and analysis. In order for a wider, more general population to receive treatment, a single drug effective over a range of genotypes is favorable. By isolating the molecular and mechanical differences between a protease variant, more precise therapies can also be created.
HIV-1 integrase nicks a conserved terminal dinucleotide at the 3’ end of HIV viral DNA. It then inserts and ligates the processed viral DNA within the chromosomal DNA of the host so that viral replication can occur. As there is only one antiretroviral drug targeted for HIV-1 integrase, resistance is frequent. In collaboration with Merck Pharmaceuticals, we aim to identify and model structural mutations in HIV-1 integrase that occur as a result of resistance. Our goal for our structure-function studies is to elucidate the mechanism of resistance for each mutation. This information will be used to design and suggest potential compounds that would inhibit the catalytic sites for a multi-resistant HIV-1 integrase.
Myelin Protein Zero
Charcot-Marie-Tooth (CMT) disease is a hereditary peripheral nervous system disorder, resulting in motor and sensory neuropathy. Our laboratory, in conjunction with the laboratory of Dr. John Kamholz, investigates the effect of a point mutation on Chromosome 1q22 as it relates to Charcot-Marie-Tooth disease Type 1 B (CMT1B). CMT1B is caused by mutations of the gene encoding Myelin Protein Zero (MPZ). MPZ, predominantly expressed in Schwann cells, is a component of the myelin sheath that surrounds, insulates, and protects peripheral nerve axons. MPZ is a transmembrane protein that inserts into the Schwann cell membrane and interacts extracellularly with the myelin sheath. Our primary focus is on the H10P mutation: the substitution of proline for histidine at position 10 is unique to human MPZ. By determining the crystal structure of the extracellular domain H10P MPZ and comparing it to the wild-type MPZ, we can analyze the molecular effects that this specific human mutation has upon the incorporation of this protein into the myelin sheath. The research of this protein contributes further to the mechanisms causing peripheral neuropathy in CMT1B patients.