Internal motion of a tryptophan residue in Streptomyces subtilisin inhibitor: deuterium nuclear magnetic resonance in solution.

Abstract

Deuterium NMR spectroscopy was used to study internal motions of a deuterium-labeled single tryptophan (Trp) residue (per subunit) of Streptomyces subtilisin inhibitor (SSI) in solution. The free inhibitor with the five ring protons of the Trp replaced with deuterons showed a narrow resonance component (56 Hz) of about one-quarter of the total intensity, in addition to the broad resonance component (about 600 Hz) at 25 degrees C, showing that it exits in an equilibrium mixture of two conformers, in one of which the tryptophan side chain is highly mobile. In analogy to the two structures of SSI found in the crystal, these two conformers were attributed to the one in which the contact between the alpha-lobe and the beta-lobe of the subunit is tight and the other in which the same contact is loose. When SSI forms a complex with subtilisin BPN', the broad component becomes invisibly broad, but the narrow component increases with even further narrowing, suggesting that the binding to the enzyme favors the "loose" conformer over the "tight" conformer.