Biology subjects

Lockbaum, G. J.

Publications and source records attributed to Lockbaum, G. J..

3 recordsLinked to original sources

bioRxiv2022

Defining the Substrate Envelope of SARS-CoV-2 Main Protease to Predict and Avoid Drug Resistance

Coronaviruses, as exemplified by SARS-CoV-2, can evolve and spread rapidly to cause severe disease morbidity and mortality. Direct acting antivirals (DAAs) are highly effective in decreasing disease burden especially when they target essential viral enzymes, such as proteases and polymerases, as demonstrated in HIV-1 and HCV and most recently SARS-CoV-2. Optimization of these DAAs through iterative structure-based drug design has been shown to be critical. Particularly, the evolutionarily conserved molecular mechanisms underlying viral replication can be leveraged to develop robust antivirals against rapidly evolving viral targets. The main protease (Mpro) of SARS-CoV-2, which is evolutionarily constrained to recognize and cleave 11 specific sites to promote viral maturation, exemplifies one such target. In this study we define the substrate envelope of Mpro by determining the molecular basis of substrate recognition, through nine high-resolution cocrystal structures of SARS-CoV-2 Mpro with the viral cleavage sites. These structures enable identification of evolutionarily vulnerable sites beyond the substrate envelope that may be susceptible to drug resistance and compromise binding of the newly developed Mpro inhibitors.

biochemistry↗

bioRxiv2019

Selection of HIV-1 for Resistance to Fourth Generation Protease Inhibitors Reveals Two Independent Pathways to High-Level Resistance

Darunavir (DRV) is exceptional among potent HIV-1 protease inhibitors (PIs) in the high drug concentrations that are achived in vivo. Little is known about the de novo resistance pathway for DRV. We have selected for resistance to high drug concentrations against ten PIs and their structural precursor DRV. Mutations accumulated through two pathways (anchored by protease mutations I50V or I84V). Small chemical changes in the inhibitor P1 side chain led to preferential use of one pathway over the other. Changes in the inhbitor P2 side chain determined differences in potency that were retained in the resistant viruses. Viral variants from the two pathways showed differential selection of compensatory mutations in Gag cleavage sites. These results reveal the high level of selective pressure that is attainable with fourth generation PIs, and how features of the inhibitor affect both the resistance pathway and the residual potency in the face of resistance.

microbiology↗

bioRxiv2018

Structural Adaptation of Darunavir Analogs Against Primary Resistance Mutations in HIV-1 Protease

HIV-1 protease is one of the prime targets of agents used in antiretroviral therapy against HIV. However, under selective pressure of protease inhibitors, primary mutations at the active site weaken inhibitor binding to confer resistance. Darunavir (DRV) is the most potent HIV-1 protease inhibitor in clinic; resistance is limited, as DRV fits well within the substrate envelope. Nevertheless, resistance is observed due to hydrophobic changes at residues including I50, V82 and I84 that line the S1/S1 pocket within the active site. Through enzyme inhibition assays and a series of 12 crystal structures, we interrogated susceptibility of DRV and two potent analogs to primary S1 mutations. The analogs had modifications at the hydrophobic P1 moiety to better occupy the unexploited space in the S1 pocket where the primary mutations were located. Considerable losses of potency were observed against protease variants with I84V and I50V mutations for all three inhibitors. The crystal structures revealed an unexpected conformational change in the flap region of I50V protease bound to the analog with the largest P1 moiety, indicating interdependency between the S1 subsite and the flap region. Collective analysis of protease-inhibitor van der Waals (vdW) interactions in the crystal structures using principle component analysis indicated I84V mutation underlying the largest variation in the vdW contacts. Interestingly, the principle components were able to distinguish inhibitor identity and relative potency solely based on vdW interactions of active site residues in the crystal structures. Our results reveal the interplay between inhibitor P1 moiety and primary S1 mutations, as well as suggesting a novel method for distinguishing the interdependence of resistance through principle component analyses.

molecular biology↗