http://www.biomedcentral.com/bmcchembiol/ The editor's pick of recent articles published by BMC Chemical Biology 2012-03-15T00:00:00Z Background: Nef is an HIV-1 accessory protein essential for viral replication and AIDS progression. Nef interacts with a multitude of host cell signaling partners, including members of the Src kinase family. Nef preferentially activates Hck, a Src-family kinase (SFK) strongly expressed in macrophages and other HIV target cells, by binding to its regulatory SH3 domain. Recently, we identified a series of kinase inhibitors that preferentially inhibit Hck in the presence of Nef. These compounds also block Nef-dependent HIV replication, validating the Nef-SFK signaling pathway as an antiretroviral drug target. Our findings also suggested that by binding to the Hck SH3 domain, Nef indirectly affects the conformation of the kinase active site to favor inhibitor association. Results: To test this hypothesis, we engineered a "gatekeeper" mutant of Hck with enhanced sensitivity to the pyrazolopyrimidine tyrosine kinase inhibitor, NaPP1. We also modified the RT loop of the Hck SH3 domain to enhance interaction of the kinase with Nef. This modification stabilized Nef:Hck interaction in solution-based kinase assays, as a way to mimic the more stable association that likely occurs at cellular membranes. Introduction of the modified RT loop rendered Hck remarkably more sensitive to activation by Nef, and led to a significant decrease in the Km for ATP as well as enhanced inhibitor potency. Conclusions: These observations suggest that stable interaction with Nef may induce Src-family kinase active site conformations amenable to selective inhibitor targeting. http://www.biomedcentral.com/1472-6769/12/1 Teodora Pene-Dumitrescu Sherry T Shu Thomas E Wales John J Alvarado Haibin Shi Purushottam Narute Jamie A Moroco Joanne I Yeh John R Engen Thomas E Smithgall BMC Chemical Biology 2012, 12:1 2012-03-15T00:00:00Z 10.1186/1472-6769-12-1 Conformational change in Hck upon Nef docking Binding of HIV-1 Nef induces conformational change in the ATP-binding site of Hck, making it more susceptible to pharmacological inhibition and thus validating the Nef-Hck signalling pathway as an antiretroviral drug target. /content/figures/1472-6769-12-1-toc.gif BMC Chemical Biology 1472-6769 12 1 2012-03-15T00:00:00Z XML