Metals affect the structure and activity of human plasminogen activator inhibitor-1. II. Binding affinity and conformational changes
Human plasminogen activator inhibitor type 1 (PAI-1) is a serine protease inhibitor with a metastable active conformation. The lifespan of the active form of PAI-1 is modulated via interaction with the plasma protein, vitronectin, and various metal ions. These metal ions fall into two categories: Type I metals, including calcium, magnesium, and manganese, stabilize PAI-1 in the absence of vitronectin, whereas Type II metals, including cobalt, copper, and nickel, destabilize PAI-1 in the absence of vitronectin, but stabilize PAI-1 in its presence. To provide a mechanistic basis for understanding the unusual modulation of PAI-1 structure and activity, the binding characteristics and conformational effects of these two types of metals were further evaluated. Steady-state binding measurements using surface plasmon resonance indicated that both active and latent PAI-1 exhibit a dissociation constant in the low micromolar range for binding to immobilized nickel. Stopped-flow measurements of approach-to-equilibrium changes in intrinsic protein fluorescence indicated that the Type I and Type II metals bind in different modes that induce distinct conformational effects on PAI-1. Changes in the observed rate constants with varying concentrations of metal allowed accurate determination of binding affinities for cobalt, nickel, and copper, yielding dissociation constants of ~40, 30, and 0.09 μM, respectively. Competition experiments that tested effects on PAI-1 stability were consistent with these measurements of affinity and indicate that copper binds tightly to PAI-1. Published by Wiley-Blackwell. © 2010 The Protein Society.
Publication Source (Journal or Book title)
Thompson, L., Goswami, S., & Peterson, C. (2011). Metals affect the structure and activity of human plasminogen activator inhibitor-1. II. Binding affinity and conformational changes. Protein Science, 20 (2), 366-378. https://doi.org/10.1002/pro.567