The molecular structure of [(Boc-Cys-Pro-Leu-Cys-OMe)Hg], 1, was studied in dimethylformamide (DMF) using X-ray absorption fine structure (XAFS), NMR, distance geometry (DG), and restrained molecular dynamics (RMD). The structure was compared with the Cys-X-Y-Cys/M(2+) (Fe2+, Zn2+; tetrahedral) sites in metalloproteins. XAFS clarified that compound 1 has a linear coordinate mercury with Hg-S bond length of 2.33 Angstrom. Approximate distances for 13 H-1-H-1 pairs in the main-chain loop were revealed by nuclear Overhauser and exchange spectroscopy (NOESY). The manner of hydrogen bonding was estimated by NMR also. The combined use of DG and RMD, as well as free molecular dynamics, based on these experimental results afforded a molecular structure that differs from that of Cys(39)-Pro-Val-Cys(41)/Fe2+ in rubredoxin (Rub). The linear Hg exerts a force up to the second residue from the C terminal in compound 1. Compound 1 shows only one hydrogen bond (Leu NH...Cys(1) S), whereas the Rub core site has two hydrogen bonds (Val NH...Cys(39) S and Cys(41) NH...Cys(39) S). The effect of linear Hg on peptide was also evaluated by phi, phi, chi torsion angles. The linear Hg distorts the C-alpha-C-beta axes of Cys(1) and Cys(4) by about 38 degrees and -18 degrees, respectively, from the stable angles, whereas the iron in the Rub core exerts a slight strain (16.7 degrees) on the C-alpha-C-beta of Cys.(41) The Leu (phi, phi) of compound 1 is found close to the beta sheet region in the Ramachandran plot, probably situated on the slope of the energy contour map. This is in contrast to the Val (phi, phi) of the Rub core site, which is situated in the alpha helix region.