Increased backbone mobility in β-barrel enhances entropy gain driving binding of N-TIMP-1 to MMP-3

The high-affinity inhibition of stromelysin 1 (MMP-3) by tissue inhibitor of metalloproteinases 1 (TIMP-1) helps control tissue remodeling and tumor development. The interaction of N-TIMP-1 with the catalytic domain of MMP-3 has been investigated by titration calorimetry and N-15 NMR. Their unfavorable enthalpy of binding of + 6.5 kcal mol(-1) is unusual among protein-protein associations, deviates from structure-based prediction, and is compensated by a net entropy increase providing at least 18 kcal mol(-1) of favorable free energy of binding at a 1 M reference state. The small heat capacity of binding agrees well with the heat capacity predicted from 65% of the surface buried on binding being polar, and suggests that the hydrophobic effect can account for only part of the entropy of binding. Using NMR, binding-induced changes in the backbone of N-TIMP-1 were checked as one possible source of conformational entropy changes. MMP binding slightly increases rigidity in some contact sites in TIMP-1 but increases mobility remotely in the otherwise rigid P-barrel core of N-TIMP-1, increasing N-15 relaxation evidence of pico- to nanosecond and micro- to millisecond fluctuations of beta-strands A-E Residual dipolar couplings suggest dynamic deviations from X-ray coordinates of the complex. These suggest that the beta-barrel has small backbone conformational fluctuations, while segments of strands betaB, betaE and betaF might experience fluctuations only in their backbone environment. This is a distinctive example of affinity between two well-structured proteins being enhanced by increased conformational entropy in the reservoir of a folding core. (C) 2003 Elsevier Science Ltd. All rights reserved.