An analytical algorithm, called SETTLE, for resetting the positions and velocities to satisfy the holonomic constraints on the rigid water model is presented. This method is still based on the Cartesian coordinate system and can be used in place of SHAKE and RATTLE. We implemented this algorithm in the SPASMS package of molecular mechanics and dynamics. Several series of molecular dynamics simulations were carried out to examine the performance of the new algorithm in comparison with the original RATTLE method. It was found that SETTLE is of higher accuracy and is faster than RATTLE with reasonable tolerances by three to nine times on a scalar machine. Furthermore, the performance improvement ranged from factors of 26 to 98 on a vector machine since the method presented is not iterative.

We present calculations of the absolute and relative binding free energies of complexation of streptavidin with biotin and its analogs by means of a thermodynamic free energy perturbation method implemented with molecular dynamics. Using the recently solved crystal structure of the streptavidin-biotin complex, biotin was mutated into a dummy molecule as well as thiobiotin and iminobiotin both in the protein and in solution. The calculated absolute binding free energy was dependent on the simulation model used. Encouragingly, the "best models" provided a reasonable semiquantitative reproduction (-20 to -22 kcal/mol) of the experimental free energy (-18.3 kcal/mol). Furthermore, the calculated results give clear insights into the binding nature of the protein-ligand complex, showing that the van der Waals energy dominates the electrostatic and hydrogen bonding energies in the binding of biotin by streptavidin. Specifically, the mutation of biotin into a dummy molecule in solution has a delta G (van der Waals) approximately -4 kcal/mol, due to the cancellation of dispersion and repulsion "cavity" effects. On the other hand, in the protein, a very small free energy price must be paid to create a cavity since one already exists and the mutation of biotin into a dummy molecule has a delta G (van der Waals) approximately 15 kcal/mol. These results are also consistent with the interpretation that the entropy increase to be expected from hydrophobic interactions from desolvation of biotin is counterbalanced by a decrease in entropy accompanying the formation of buried hydrogen bonds, which have been derived from the apparently conflicting experimental data. They provide an alternative interpretation of the reason for the extremely high affinity of the biotin-streptavidin interaction than that recently proposed by Weber et al. (J. Am. Chem. Soc. 114:3197, 1992). In the case of the relative binding free energies, the calculated values of 3.8 +/- 0.6 and 7.2 +/- 0.6 kcal/mol compare well with the experimental values of 3.6 and 6.2 kcal/mol for the perturbation of biotin to thiobiotin and iminobiotin, respectively in the related protein avidin. The calculations indicate that desolvation of the ligand is important in understanding the relative affinity of the ligands with the protein. The above successful simulations suggest that the molecular dynamics/free energy perturbation method is useful for understanding the energetic features affecting the binding between proteins and ligands, since it is generally difficult to determine these factors unambiguously by experiment.(ABSTRACT TRUNCATED AT 400 WORDS)

Free energy perturbation methods using molecular dynamics have been used to calculate the absolute free energy of association of two ligand-protein complexes. The calculations reproduce the snificantly more negative free energy of ation of biotin to streptavidin, compared to N-L-acetyltryptophanamlide/a-chymotrypsln. This difference in free energy ofassocition is due to van der Waals/dlspersion effects in the nearly ideally preformed cavity that streptavidin presents to biotin, which involves four ryptophan residues.One of the exciting developments in computer modeling of complex molecules in solution has been the capability to calculate relative free energies of association of these molecules and to relate these values to experiment (1, 2). This development has been catalyzed by methodological advances (3, 4) and increased computer capabilities. In favorable cases, relative free energies of association within 1 kcal/mol (1 cal = 4.184 J) of experiment have been achieved (2). In such cases, the calculations could be of use in experimental ligand design. However, inaccuracies in molecular mechanical force fields and representation of the system and, even more importantly, limitations in one's ability to completely sample the relevant regions of conformational space, have restricted the number of systems to which such free energy calculations could be applied to give chemical accuracy (5,6).Nonetheless, such free energy calculations can be very valuable and interesting even when such accuracy is not achieved, because mechanistic insight into noncovalent association in general and protein-ligand design in particular can be extracted from them (7,8 (14) have had success with this approach for nucleic acid bases in organic solvents, and Lee et al. (15) have calculated the absolute free energy of association of phosphorylcholine analogs to an immunoglobulin, but no one has carried out such a large and dramatic change as in the biotin-streptavidin association studied by Miyamoto and Koilman (9). In that paper, either all of biotin or all of biotin but the terminal COj group were mutated to dummy atoms both in water and in the protein; in either case, a AG for association in the range of -20 kcal/mol could be calculated, in good agreement with experiment. Given the approximations in that study (neglect of any changes in intramolecular energies of biotin free and bound and underestimate of translational/rotational entropy losses due to the use of hydrogen bond restraints) and the difficulty in precisely estimating the magnitude of the errors, we turned to another ligand-protein complex with much lower affinity, with a ligand of size comparable to biotin to use as a control. The results of that calculation, presented here along with the results of the biotin-streptavidin calculation, enable fundamental insights into the nature ofligand-protein associations. METHODSWe used the free energy perturbation method to calculate the binding free energy of complexation. This method uses an easily derived equation from class...

A series of imidazole-5-carboxylic acids bearing alkyl, alkenyl, and hydroxyalkyl substituents at the 4-position and their related compounds were prepared and evaluated for their antagonistic activities to the angiotensin II (AII) receptor. Among them, the 4-(1-hydroxyalkyl)-imidazole derivatives had strong binding affinity to the AII receptor and potently inhibited the AII-induced pressor response by intravenous administration. Various esters of these acids showed potent and long-lasting antagonistic activity by oral administration. The most promising compounds were (5-methyl-2-oxo-1,3-dioxol-4-yl)methyl (CS-866) and (pivaloyloxy)-methyl esters of 4-(1-hydroxy-1-methylethyl)-2-propyl-1-[(2'-1H-tetrazol-5- ylbiphenyl-4-yl)-methyl]imidazole-5-carboxylic acid (26c). A study involving stereochemical comparison of 26c with the acetylated C-terminal pentapeptide of AII was also undertaken.

Background and Aim: Prophylactic clipping has been widely used to prevent post-procedural bleeding in colon polypctomy. However, its efficiency has not been confirmed and there is no consensus on the usefulness of prophylactic clipping. The aim of the present study was to evaluate the preventive effect of prophylactic clipping on post-polypectomy bleeding.Methods: A multicenter randomized controlled study was conducted from January 2012 to July 2013 in Japan. Patients who had polyps <2 cm in diameter were divided into a clipping group and a non-clipping group by cluster randomization. After endoscopic polypectomy, patients allocated to the clipping group underwent prophylactic clipping, whereas the procedure was completed without clipping in patients allocated to the nonclipping group. Occurrence of post-polypectomy bleeding was compared between the two groups.Results: Seven hospitals participated in this study. A total of 3365 polyps in 1499 patients were evaluated. The clipping group consisted of 1636 polyps in 752 patients, and the non-clipping group consisted of 1729 polyps in 747 patients. Postpolypectomy bleeding occurred in 1.10% (18/1636) of the cases in the clipping group, and in 0.87% (15/1729) of those in the nonclipping group. The difference was -0.22% (95% confidence interval [CI]: -0.96, 0.53). Upper limit of the 95% CI was lower than the noninferiority margin (1.5%), and we could thus prove non-inferiority of non-clipping against clipping.Conclusion: Prophylactic clipping is not necessary to prevent post-polypectomy bleeding for polyps <2 cm in diameter.