Molybdate inhibits hsp90, induces structural changes in its C-terminal domain, and alters its interactions with substrates

To examine the biochemical mechanism by which hsp90 exerts its essential po sitive function on certain signal transduction proteins, we characterized t he effects of molybdate and geldanamycin on hsp90 function and structure. M olybdate inhibited hsp90-mediated p56(lck) biogenesis and luciferase renatu ration while enforcing salt-stable interactions with these substrates. Moly bdate also reduced the amount of free hsp90 present in cell lysates, inhibi ted hsp90's ability to bind geldanamycin, and induced resistance to proteol ysis at a specific region within the C-terminal domain of hsp90. In contras t, the hsp90 inhibitor geldanamycin prevented hsp90 from assuming natural o r molybdate-induced conformations that allow salt-stable interactions with substrates. When these compounds were applied sequentially, the order of ad dition determined the effects observed, indicating that these agents had op posing effects on hsp90. We conclude that a specific region within the C-te rminal domain of hsp90 (near residue 600) determines the mode by which hsp9 0 interacts with substrates and that the ability of hsp90 to cycle between alternative modes of interaction is obligatory for hsp90 function.