How the Hinge Region Affects Interactions between the Catalytic and β-Propeller Domains in Oligopeptidase B

Abstract

In order to elucidate the effect of modification of the hinge region on structural polymorphism associated with conformational transitions, structural studies of hinge-modified oligopeptidase B from Serratia proteamaculans (SpOpBmod) in the crystalline state and solution were carried out. A new crystal structure of SpOpBmod in the intermediate conformation was obtained, and a molecular model of SpOpBmod in the open conformation was created using a combination of small-angle X-ray scattering with MD simulations. The improved electron density of the mobile H-loop carrying the catalytic H652 distinguished the obtained crystal structure from that which was previously reported. Good electron density in this region was previously found only in the inhibitor-bound SpOpBmod structure, in which one of the inhibitor molecules was covalently bound to H652. Comparison of the above structures of free and inhibitor-bound enzymes showed that both tertiary folds are the result of the internal conformational dynamics of SpOpBmod, which were captured by inhibitor binding. Comparison of the SpOpBmod structures with the structures of the same enzyme with a native hinge peptide made it possible to establish the influence of hinge modification on the rearrangement of the interdomain interface during conformational transitions. The above analysis also used models of native and hinge-modified enzymes in open conformations. We found that the interdomain interface observed in the crystal structures of hinge-modified enzymes could be considered an extreme version of the H-loop arrangement, in which closure of the domains does not lead to the assembly of the catalytic triad, whereas the intermediate conformation observed in the structure of the enzyme with the native hinge sequence illustrates a productive transition to the catalytically active closed conformation.