Semi-quantification and Potency Verification of the HIV Protease Inhibitor Based on the Matrix-Capsid Protein Immobilized Nickel (II)/NTA-Tol/Graphene Oxide/SPCE Electrochemical Biosensor.

Human immunodeficiency virus (HIV) causing acquired immune deficiency syndrome (AIDS) is still a global issue. Long-term drug treatment and nonadherence to medication increase the spread of drug-resistant HIV strains. Therefore, the identification of new lead compounds is being investigated and is highly desirable. Nevertheless, a process generally necessitates a significant budget and human resources. In this study, a simple biosensor platform for semi-quantification and verification of the potency of HIV protease inhibitors (PIs) based on electrochemically detecting the cleavage activity of the HIV-1 subtype C-PR (C-SA HIV-1 PR) was proposed. An electrochemical biosensor was fabricated by immobilizing His6-matrix-capsid (H6MA-CA) on the electrode surface via the chelation to Ni2+-nitrilotriacetic acid (NTA) functionalized GO. The functional groups and the characteristics of modified screen-printed carbon electrodes (SPCE) were characterized by Fourier transform infrared (FTIR) spectroscopy, scanning electron microscopy (SEM), and energy-dispersive X-ray spectroscopy (EDS). C-SA HIV-1 PR activity and the effect of PIs were validated by recording changes in electrical current signals of the ferri/ferrocyanide redox probe. The detection of PIs, i.e., lopinavir (LPV) and indinavir (IDV), toward the HIV protease was confirmed by the decrease in the current signals in a dose-dependent manner. In addition, our developed biosensor demonstrates the ability to distinguish the potency of two PIs to inhibit C-SA HIV-1 PR activities. We anticipated that this low-cost electrochemical biosensor would increase the efficiency of the lead compound screening process and accelerate the discovery and development of new HIV drugs.

Understanding Drug Resistance of Wild-Type and L38HL Insertion Mutant of HIV-1 C Protease to Saquinavir.

Acquired immunodeficiency syndrome (AIDS) is one of the most challenging infectious diseases to treat on a global scale. Understanding the mechanisms underlying the development of drug resistance is necessary for novel therapeutics. HIV subtype C is known to harbor mutations at critical positions of HIV aspartic protease compared to HIV subtype B, which affects the binding affinity. Recently, a novel double-insertion mutation at codon 38 (L38HL) was characterized in HIV subtype C protease, whose effects on the interaction with protease inhibitors are hitherto unknown. In this study, the potential of L38HL double-insertion in HIV subtype C protease to induce a drug resistance phenotype towards the protease inhibitor, Saquinavir (SQV), was probed using various computational techniques, such as molecular dynamics simulations, binding free energy calculations, local conformational changes and principal component analysis. The results indicate that the L38HL mutation exhibits an increase in flexibility at the hinge and flap regions with a decrease in the binding affinity of SQV in comparison with wild-type HIV protease C. Further, we observed a wide opening at the binding site in the L38HL variant due to an alteration in flap dynamics, leading to a decrease in interactions with the binding site of the mutant protease. It is supported by an altered direction of motion of flap residues in the L38HL variant compared with the wild-type. These results provide deep insights into understanding the potential drug resistance phenotype in infected individuals.