Effects of trace metal compounds on HIV-1 reverse transcriptase: an in vitro study
Biol Trace Elem Res 1999 May;68(2):107-19 (ISSN: 0163-4984)
Rivas CI; Vera JC; Guaiquil VH; Velasquez FV; Borquez-Ojeda
OA; Carcamo JG; Concha II; Golde DW Program in Molecular Pharmacology
and Therapeutics, Memorial Sloan-Kettering Cancer Center, New York, New
York 10021, USA.
The effect of 44 different metal ions (Ag+, Al3+, As(O-)2, Au3+, Ba2+,
Be2+, Bi3+, Cd2+, Ce3+, CO2+, Cr(O2-)4, Cr3+, Cs+, Cu2+, Fe3+, Fe2+, Ga3+,
Ge4+, Hg2+, Ir4+, La3+, Li+, Mn2+, MO6+, Ni2+, OS4+, Pb2+, Pt4+, Rb+,
Rh3+, Sb5+, Se(O2-)4, Se(O2-)3, Sn2+, Sr2+, Th4+, T1+, U(O2+)2, V(O-)3,
VO2+, W(O2-)4, Y3+, Zn2+, and Zr4+) on the activity of the reverse transcriptase
(RT) of the human immunodeficiency virus (HIV-1) was investigated in vitro.
For this study, the RT activity assay was carried out by means of an enzyme-linked
immunosorbent assay (ELISA) kit, using the template/primer hybrid poly(A)
oligo(dT)15, which required some modifications: (1) possible interfering
metal chelators (such as EDTA) in the original lysis buffer were avoided,
and a new buffer (50 mM Tris-NO3, pH 7.8) was used throughout; (2) an
amount of 2 ng of RT per well was considered to be optimal after checking
the linearity of the reaction with increasing amounts of enzyme; (3) an
incubation temperature of 37 degrees C and an incubation time of 1 h were
chosen after preliminary studies in a wide range of temperature and time.
At an incubation temperature > or = 40 degrees C, there was a dramatic
loss of enzymatic activity. In addition, when RT alone was preincubated
for 1 h at 5 degrees C, 25 degrees C, and 37 degrees C, there was a large
(83%) loss of activity at 37 C as compared to that at 5 degrees C. These
results are indicative of enzyme thermolability, which is higher in the
absence of substrates. The effect of metal ions on RT activity was tested
using two different metal salt concentrations (10(-4) M and 10(-5) M).
Under such experimental conditions, the presence of five metal ions (Pt4+,
Ag+, Rh3+, Zn2+, and Hg2+) decreased the RT activity in a dose-response
fashion. The observed order of effectiveness with respect to inhibition
was Pt4+ > Ag+ > Rh3+ > Zn2+ = Hg2+. Estimated mean inhibitory
concentrations (IC50) were 7.8 microM for (NH4)2PtCl6, 14.1 microM for
AgNO3, 46.8 microM for RhCl3, 53.7 microM for Zn(SO)4, and 56.2 microM
for Hg(NO3)2. Because these data are of the same order of magnitude as
the corresponding values related to other RT inhibitors used in anti-AIDS
therapy, metal compounds or their derivatives could give an interesting
contribution in the development of new RT inhibitors for clinical use.