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The solution conformations of the protein hirudin have been investigated by the combined use of distance geometry and restrained molecular dynamics calculations. The basis for the structure determination comprised 359 approximate interproton distance restraints and 10 4 backbone torsion angle restraints derived from n.m.r. measurements. It is shown that hirudin is composed of three domains: a central core made up of residues 3-30, 37-46 and 56-57; a protruding 'finger' (residues 31-36) consisting of the tip of an antiparaliel 3 sheet, and an exposed loop (residues 47-55). The structure of each individual domain is relatively well defined with average backbone atomic r.m.s. differences of <2 A between the final seven converged restrained dynamic structures and the mean structure obtained by averaging their coordinates. The orientation of the two minor domains relative to the central core, however, could not be determined as no long-range (I i-jI >5) interdomain proton-proton contacts could be observed in the two-dimensional nuclear Overhauser enhancement spectra. From the restrained molecular dynamics calculations it appears that the two minor domains exhibit large rigid-body motions relative to the central core.

The conformation of the polypeptide thymosin beta 4 in solutions of 60% (v/v) trifluoroethanol-d3 and 50% (v/v) hexafluoroisopropyl-d2 alcohol in water is investigated by nuclear magnetic resonance (NMR) spectroscopy. Under these conditions thymosin beta 4 adopts an ordered structure. By use of a combination of two-dimensional NMR techniques, the 1H NMR spectrum of thymosin beta 4 is assigned. A set of 180 approximate interproton distance constraints is derived from nuclear Overhauser enhancement (NOE) measurements. These, together with 33 phi constraints obtained for JNH alpha coupling data and the 23 psi dihedral angles identified on the basis of the pattern of short-range NOEs, form the basis of a three-dimensional structure determination by dynamical simulated annealing. The calculations are carried out starting from three initial structures, an alpha-helix, an extended beta-strand, and a mixed alpha/beta structure. Ten independent structures are computed from each starting structure by using different random number seeds for the assignments of the initial velocities. All 30 calculated structures satisfy the experimental constraints, display very small deviations from idealized covalent geometry, and possess good nonbonded contacts. Analysis of the 30 converged structures indicates that there are two helical regions extending from residues 4-16 and from residues 30-40, which are well defined both in terms of atomic root mean square differences and backbone torsion angles. For the two helical regions individually the average backbone rms difference between all pairs of structures is approximately 2 A. The two helices exhibit typical amino acid preferences for specific locations at the ends of helices.(ABSTRACT TRUNCATED AT 250 WORDS)

The polypeptide fold of the 79-residue globular domain of chicken histone H5 (GH5) in solution has been determined by the combined use of distance geometry and restrained molecular dynamics calculations. The structure determination is based on 307 approximate interproton distance restraints derived from n.m.r. measurements. The structure is composed of a core made up of residues 3-18, 23-34, 37-60 and 71-79, and two loops comprising residues 19-22 and 61-70. The structure of the core is well defined with an average backbone atomic r.m.s. difference of 2.3 0.3 A between the final eight converged restrained dynamics structures and the mean structure obtained by averaging their coordinates best fitted to the core residues. The two loops are also well defined locally but their orientation with respect to the core could not be determined as no long range (li-jl > 5) protonproton contacts could be observed between the loop and core residues in the two-dimensional nuclear Overhauser enhancement spectra. The structure of the core is dominated by three helices and has a similar fold to the Cterminal DNA binding domain of the cAMP receptor protein.

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