Electrochemical probing of HIV enzymes using ferrocene-conjugated peptides on surfaces.
One of the current pathways to develop inhibitors that target different steps in the life cycle of the human immunodeficiency virus (HIV) is blocking the function of the HIV-related proteins such as HIV-1 integrase (HIV-1 IN), HIV-1 reverse transcriptase (HIV-1 RT) and HIV-1 protease (HIV-1 PR), which are essential proteins that control the ability of HIV to infect cells, produce new copies of the virus, or cause disease. We have demonstrated for the first time the detection of these enzymes at nanomolar levels using ferrocene (Fc)-conjugated peptides on gold microelectrodes. The interaction between the Fc-conjugated peptides and the enzymes was studied by cyclic voltammetry. As the protein concentration increased, the electrochemical behaviour of the surface-bound Fc- bioconjugate changed, indicating that HIV protein was binding to the peptide film and encapsulating the Fc redox center on the surface. The electrochemical responses shifted to higher potentials and decreased in the current intensity, as the concentrations of the HIV-1 enzymes increased. The optimization studies were performed by changing the pH and NaCl concentration. Control experiments involved the exposure of the Fc-conjugated peptides with all the enzymes. This general procedure can be readily applied in the future to the multiplexed detection of several HIV-related proteins, as well as the high-throughput screening of candidate inhibitors for AIDS therapy.
Kerman
Kagan
K
Department of Chemistry, The University of Western Ontario, 1265 Richmond Street, London, ON, Canada N6A 5B7.
Kraatz
Heinz-Bernhard
HB
D001665:Binding Sites; D055664:Electrochemical Techniques / Q000295:instrumentation* / Q000379:methods; D005296:Ferrous Compounds / Q000737:chemistry*; D015497:HIV-1 / Q000201:enzymology*; D054298:Human Immunodeficiency Virus Proteins / Q000737:chemistry*; D000075163:Metallocenes; D036103:Nanotechnology; D010455:Peptides / Q000737:chemistry*; D011485:Protein Binding; D013499:Surface Properties
D005296:Ferrous Compounds; D054298:Human Immunodeficiency Virus Proteins; D000075163:Metallocenes; D010455:Peptides; C004998:ferrocene