Quantitative analysis of the interactions between HIV-1 integrase and retroviral reverse transcriptases.
The RT (reverse transcriptase) of HIV-1 interacts with HIV-1 IN (integrase) and inhibits its enzymatic activities. However, the molecular mechanisms underling these interactions are not well understood. In order to study these mechanisms, we have analysed the interactions of HIV-1 IN with HIV-1 RT and with two other related RTs: those of HIV-2 and MLV (murine-leukaemia virus). All three RTs inhibited HIV-1 IN, albeit to a different extent, suggesting a common site of binding that could be slightly modified for each one of the studied RTs. Using surface plasmon resonance technology, which monitors direct protein-protein interactions, we performed kinetic analyses of the binding of HIV-1 IN to these three RTs and observed interesting binding patterns. The interaction of HIV-1 RT with HIV-1 IN was unique and followed a two-state reaction model. According to this model, the initial IN-RT complex formation was followed by a conformational change in the complex that led to an elevation of the total affinity between these two proteins. In contrast, HIV-2 and MLV RTs interacted with IN in a simple bi-molecular manner, without any apparent secondary conformational changes. Interestingly, HIV-1 and HIV-2 RTs were the most efficient inhibitors of HIV-1 IN activity, whereas HIV-1 and MLV RTs showed the highest affinity towards HIV-1 IN. These modes of direct protein interactions, along with the apparent rate constants calculated and the correlations of the interaction kinetics with the capacity of the RTs to inhibit IN activities, are all discussed.
Herschhorn
Alon
A
Department of Cell and Developmental Biology, Sackler School of Medicine, Tel Aviv University, Tel Aviv 69978, Israel.
Oz-Gleenberg
Iris
I
Hizi
Amnon
A
D004277:DNA, Single-Stranded / Q000378:metabolism; D004800:Enzymes, Immobilized / Q000378:metabolism; D019427:HIV Integrase / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D054303:HIV Reverse Transcriptase / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D009052:Leukemia Virus, Murine / Q000201:enzymology / Q000235:genetics; D008954:Models, Biological; D011485:Protein Binding / Q000502:physiology; D012194:RNA-Directed DNA Polymerase / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D012191:Retroviridae Proteins / Q000032:analysis / Q000378:metabolism; D013379:Substrate Specificity
D004277:DNA, Single-Stranded; D004800:Enzymes, Immobilized; D012191:Retroviridae Proteins; D019427:HIV Integrase; C514824:reverse transcriptase, Human immunodeficiency virus 1; C072416:reverse transcriptase, Human immunodeficiency virus 2; D054303:HIV Reverse Transcriptase; D012194:RNA-Directed DNA Polymerase; C514801:p31 integrase protein, Human immunodeficiency virus 1