H++: a server for estimating pKas and adding missing hydrogens to macromolecules

Abstract

The structure and function of macromolecules depend critically on the ionization (protonation) states of their acidic and basic groups. A number of existing practical methods predict protonation equilibrium pK constants of macromolecules based upon their atomic resolution Protein Data Bank (PDB) structures; the calculations are often performed within the framework of the continuum electrostatics model. Unfortunately, these methodologies are complex, involve multiple steps and require considerable investment of effort. Our web server provides access to a tool that automates this process, allowing both experts and novices to quickly obtain estimates of pKs as well as other related characteristics of biomolecules such as isoelectric points, titration curves and energies of protonation microstates. Protons are added to the input structure according to the calculated ionization states of its titratable groups at the user-specified pH; the output is in the PQR (PDB + charges + radii) format. In addition, corresponding coordinate and topology files are generated in the format supported by the molecular modeling package AMBER. The server is intended for a broad community of biochemists, molecular modelers, structural biologists and drug designers; it can also be used as an educational tool in biochemistry courses.

Novel cinnamic acid/4-aminoquinoline conjugates bearing non-proteinogenic amino acids: Towards the development of potential dual action antimalarials

“…The Xray structure of FP2 and FP3 (PDB code: 3BPF [7] and 3BWK [16], respectively) were used and prepared for docking. The original ligand, ions and solvent molecules were removed and the proteins were protonated using the Hþþ server [52] assuming a pH of 5.5 and a salinity of 0.15 mol/L. The proteins were then minimized with the AMBER program [53] by 500 steps of steepest descent followed by 2000 steps of conjugate gradient to remove bad contacts using a generalized-Born solvent model.…”

Study of the interaction of Huperzia saururus Lycopodium alkaloids with the acetylcholinesterase enzyme

“…The coordinates of the missing residues (Pro259, Gly260, Gly261, Thr262 and Gly263) were added with the side chain prediction tools included in Maestro. [55] The protonation states of the amino acids in the protein were assigned with the H++ server, [56,57] and the ones of the residues in the catalytic gorge (His447, Glu334, Glu202 and Asp74) on the basis of previous representative work by McCammon et al [58] For A3 and A4.2, the search that provided the best exploration was found to be the combination of the genetic algorithm and a local search refinement (GA-LS). The search space was defined using AutoGrid, and the same grid was used for all the simulations with the three programs.…”

Toward the discovery of inhibitors of babesipain-1, a Babesia bigemina cysteine protease: in vitro evaluation, homology modeling and molecular docking studies

“…Briefly, the proteins were protonated using the H?? server [25] assuming a pH of 5.5 and a salinity of 0.15 mol/L. The proteins were then minimized with the AMBER 11 program [26] by 500 steps of steepest descent, followed by 2,000 steps of conjugate gradient to remove bad contacts using a generalized-Born solvent model.…”