Modeling techniques for analysing conformational transitions in hemocyanins by small-angle scattering of X-rays and neutrons

摘要

Small-angle scattering (SAS) of X-rays (SAXS) or neutrons (SANS) enables investigation of the tertiary and quaternary structure of proteins in solution. Because ii is comparatively easy to add or remove chemical compounds from the protein solution, SAS is especially suited to investigate changes in the molecular structure caused by the binding of substrates or effectors. Although the resolution of the data is only one-dimensional and is limited in practice to above 1 nm, SAS is often applied successfully to large biomolecules (Svergun and Koch, 2002). One major challenge of the method is the reconstruction of three-dimensional models from one-dimensional data (Chacon et al., 1998; Svergun, 1999). Due to the low information content of the SAS data there is a considerable risk of creating an ambiguous model of the molecule, or to over-fit the data. Here, we present two methodical approaches with SAS to obtain information on the large cooperative hemocyanins and their structural transitions when binding the ligand dioxygen or allosteric effectors: (a) A rigid body modelling was developed and used to reconstruct n X 6-meric hemocyanins and to investigate different conformations of the oxy- and deoxystates (Hartmann and Decker, 2002) and conformation changes induced by effectors such as lactate (Hartmann et al., 200 lb). (b) A newly developed Monte-Carlo annealing algorithm (MCSAS) was applied to SAXS and SANS data to determine the conformations of the molluscan hemocyanin KLH1 in the oxy- and the deoxystate (Hartmann et al., 2001a; Hartmann and Decker, 2003; Hartmann et al., 2004).