Impedance method for detecting HIV-1 protease and screening for its inhibitors using ferrocene-peptide conjugate/Au nanoparticle/single-walled carbon nanotube modified electrode

Khaled A. Mahmoud, John H.T. Luong

Research output: Contribution to journalArticle

77 Citations (Scopus)

Abstract

A highly sensitive screening assay based on electrochemical impedance spectroscopy (EIS) has been developed for detecting HIV-1 protease (PR) and subsequent evaluation of its corresponding inhibitors at picomolar levels. The assay format was based on the immobilization of the thiol terminated ferrocene(Fc)-pepstatin conjugate on a single-walled carbon nanotube/gold nanoparticle (SWCNT/AuNP) modified gold electrode. The alteration of the interfacial properties of electrodes upon HIV-1 PR and Fc-pepstatin conjugate interaction was traced by EIS. On the basis of the charge transfer resistance data obtained and using a mixed kinetic and diffusion model, this procedure was capable of detecting picomolar HIV-1 PR owing to the specific binding of this enzyme to Fc modified pepstatin. A competitive inhibition assay format was then performed using four potent HIV-1 PR inhibitors. The estimated inhibition constant (Ki) attested that lopinavir/ritonavir (Ki = 20 ± 3 pM) and saquinavir (Ki = 57 ± 8 pM) even at 10 pM competed strongly with pepstatin for effective binding to HIV-1 PR. Indinavir (Ki = 630 ± 22 pM) only competed well with pepstatin at a much higher concentration (1 nM). No significant inhibitory effect was observed for the fosamprenavir (Ki = 11 ± 0.5 nM) as expected from this pro-drug. Such results agreed well with the values reported in the literature. This assay format is a definite asset for the expedited development of effective HIV-1 PR inhibitors with low molecular weights.

Original languageEnglish
Pages (from-to)7056-7062
Number of pages7
JournalAnalytical Chemistry
Volume80
Issue number18
DOIs
Publication statusPublished - 15 Sep 2008

ASJC Scopus subject areas

  • Analytical Chemistry

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