Structures of active and latent PAI-1: A possible stabilizing role for chloride ions

Serpins exhibit a range of physiological roles and can contribute to certai n disease states dependent on their various conformations. Understanding th e mechanisms of the large-scale conformational reorganizations of serpins m ay lead to a better understanding of their roles in various cardiovascular diseases. We have studied the serpin, plasminogen activator inhibitor 1 (PA I-1), in both the active and the latent state and found that anionic halide ions may play a role in the active-to-latent structural transition. Crysta llographic analysis of a stable mutant form of active PAI-1 identified an a nion-binding site between the central beta-sheet and a small surface domain . A chloride ion was modeled in this site, and its identity was confirmed b y soaking crystals in a bromide-containing solution and calculating a cryst allographic difference map. The anion thus located forms a 4-fold ligated l inchpin that tethers the surface domain to the central beta-sheet into whic h the reactive center loop must insert during the active-to-latent transiti on. Timecourse experiments measuring active PAI-1 stability in the presence of various halide ions showed a clear trend for stabilization of the activ e form with F- > Cl- > Br- >> I-. We propose that the "stickiness" of this pin (i.e., the electronegativity of the anion) contributes to the energetic s of the active-to-latent transition in the PAI-1 serpin.