From Protein Structure to Function with Bioinformatics.pdf

222 M.B. Kubitzki et al.Fig. 9.2 Nucleoplasmic (left) and cytoplasmic (right) form of Cse1p. In the nucleoplasmic form,Cse1p is bound to RanGTP and importin-α (both not shown) and adopts a superhelical structure.After dissociation in the cytoplasm, Cse1p undergoes a large conformational change and forms aring conformation that occludes the RanGTP binding site and prevents reassociation of the complex.The structures are coloured in a spectrum from blue (N-terminus) to red (C-terminus)and thus, presumably, no significant energy barrier had to be overcome to reachthe closed conformation. When simulations are started from a free energy minimum,which is usually the case, the accessible time scales are often too short toovercome higher energy barriers and, thus, to observe functionally relevant conformationaltransitions. This is known as the “sampling problem” and is a generalproblem for MD simulations.9.1.2.2 LysozymeMD simulations of bacteriophage T4-lysozyme (T4L), an enzyme which is sixtimes smaller than Cse1p, impressively illustrate this sampling problem for relativelylong MD trajectories. T4L has been extensively studied with X-ray crystallography(Faber and Matthews 1990; Kuroki et al. 1993) and, since it has beencrystallized in many different conformations, represents one of the rare caseswhere information about functionally relevant modes can be directly obtained atatomic resolution from experimental data (Zhang et al. 1995; de Groot et al.1998). The domain character of this enzyme is very pronounced (Matthews andRemington 1974) and from the differences between crystallographic structuresof various mutants of T4L it has been suggested that a hinge-bending mode ofT4L (Fig. 9.3) is an intrinsic property of the molecule (Dixon et al. 1992).Moreover, the domain fluctuations are predicted to be essential for the functionof the enzyme, allowing the substrate to enter and the products to leave theactive site in the open configuration, with the closed state presumably requiredfor catalysis.