scholarly journals HIV Reverse Transcriptase Pre-Steady-State Kinetic Analysis of Chain Terminators and Translocation Inhibitors Reveals Interactions between Magnesium and Nucleotide 3′-OH

ACS Omega ◽  
2021 ◽  
Author(s):  
Christopher R. Dilmore ◽  
Jeffrey J. DeStefano
Keyword(s):  
Steady State ◽  
Reverse Transcriptase ◽  
Kinetic Analysis ◽  
Hiv Reverse Transcriptase ◽  
Steady State Kinetic ◽  
State Kinetic ◽  
Chain Terminators

scholarly journals Pre-steady state kinetic analysis of cyclobutyl derivatives of 2′-deoxyadenosine 5′-triphosphate as inhibitors of HIV-1 reverse transcriptase

2012 ◽  
Vol 22 (12) ◽  
pp. 4064-4067 ◽  
Author(s):  
Jiae Kim ◽  
Ligong Wang ◽  
Yongfeng Li ◽  
Kimberlynne D. Becnel ◽  
Kathleen M. Frey ◽  
...  
Keyword(s):  
Steady State ◽  
Reverse Transcriptase ◽  
Kinetic Analysis ◽  
Steady State Kinetic ◽  
State Kinetic ◽  
Derivatives Of ◽  
Hiv 1

scholarly journals HIV reverse transcriptase pre-steady-state kinetic analysis of chain terminators and translocation inhibitors reveals interactions between magnesium and nucleotide 3’-OH

2020 ◽  
Author(s):  
Christopher R. Dilmore ◽  
Jeffrey J. DeStefano
Keyword(s):  
Steady State ◽  
Reverse Transcriptase ◽  
Hiv Reverse Transcriptase ◽  
Oh Groups ◽  
Steady State Kinetics ◽  
Steady State Kinetic ◽  
Sugar Ring ◽  
Group 2 ◽  
Equilibrium Dissociation ◽  
State Kinetic

AbstractDeoxythymidine triphosphate analogs with various 3’ groups (-OH (dTTP), -H, -N3, -NH2, -F, -O- CH3, and no group (2′,3′-didehydro-2′,3′-dideoxythymidine triphosphate (d4TTP)), and those retaining the 3’-OH but with 4’ additions (4’-C-methyl, 4’-C-ethyl) or sugar ring modifications (D-carba dTTP) were evaluated using pre-steady-state kinetics in low (0.5 mM) and high (6 mM) Mg2+ with HIV reverse transcriptase (RT). All analogs showed diminished incorporation compared to dTTP ranging from about 2-fold (3’-H, -N3, and d4TTP with 6 mM Mg2+) to >10-fold (3’-NH2 and 3’-F with 0.5 mM Mg2+). The exception was 3’-O-CH3 dTTP which was incorporate profoundly more slowly than other analogs. The incorporation rate (k) using 5 µM dTTP and 0.5 mM (free) Mg2+ was modestly slower (1.6-fold) than with 6 mM Mg2+, while analogs with 3’ modifications were incorporated more slowly (2.8-5.1-fold) in 0.5 mM Mg2+. In contrast, 4’-C-methyl and D-carb, which retain the 3’-OH, were not significantly affected by Mg2+. Consistent with the above results, analogs with 3’ modifications were better inhibitors with 6 mM vs. 0.5 mM Mg2+, in primer extension reactions on a long template. Equilibrium dissociation constant (Kd) and kpol determinations for dTTP and analogs lacking a 3’-OH indicated that low Mg2+ caused a several-fold greater reduction in kpol with the analogs but had little effect on Kd. Overall, results emphasize the importance of as yet undefined interactions between Mg2+ and the 3’-OH and indicate that inhibitors with 3’-OH groups may have an advantage in a physiological setting where the concentration of free Mg2+ is low.

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