Interaction Entropy for Computational Alanine Scanning.

The theoretical calculation of protein-protein binding free energy is a grand challenge in computational biology. Accurate prediction of critical residues along with their specific and quantitative contributions to protein-protein binding free energy is extremely helpful to reveal binding mechanisms and identify drug-like molecules that alter protein-protein interactions. In this paper, we propose an interaction entropy approach combined with the molecular mechanics/generalized Born surface area (MM/GBSA) method for solvation to compute residue-specific protein-protein binding free energy. In the current approach, the entropic loss in binding free energy of individual residues is explicitly computed from moledular dynamics (MD) simulation by using the interaction entropy method. In this approach the entropic contribution to binding free energy is determined from fluctuation of the interaction in MD simulation. Studies for an extensive set of realistic protein-protein interaction systems showed that by including the entropic contribution, the computed residue-specific binding free energies are in better agreement with the corresponding experimental data.

Molecular dynamics study of DNA binding by INT-DBD under a polarized force field.

The DNA binding domain of transposon Tn916 integrase (INT-DBD) binds to DNA target site by positioning the face of a three-stranded antiparallel ß-sheet within the major groove. As the negatively charged DNA directly interacts with the positively charged residues (such as Arg and Lys) of INT-DBD, the electrostatic interaction is expected to play an important role in the dynamical stability of the protein-DNA binding complex. In the current work, the combined use of quantum-based polarized protein-specific charge (PPC) for protein and polarized nucleic acid-specific charge (PNC) for DNA were employed in molecular dynamics simulation to study the interaction dynamics between INT-DBD and DNA. Our study shows that the protein-DNA structure is stabilized by polarization and the calculated protein-DNA binding free energy is in good agreement with the experimental data. Furthermore, our study revealed a positive correlation between the measured binding energy difference in alanine mutation and the occupancy of the corresponding residue's hydrogen bond. This correlation relation directly relates the contribution of a specific residue to protein-DNA binding energy to the strength of the hydrogen bond formed between the specific residue and DNA.

Effect of interprotein polarization on protein-protein binding energy.

Molecular dynamics simulation in explicit water for the binding of the benchmark barnase-barstar complex was carried out to investigate the effect polarization of interprotein hydrogen bonds on its binding free energy. Our study is based on the AMBER force field but with polarized atomic charges derived from fragment quantum mechanical calculation for the protein complex. The quantum-derived atomic charges include the effect of polarization of interprotein hydrogen bonds, which was absent in the standard force fields that were used in previous theoretical calculations of barnase-barstar binding energy. This study shows that this polarization effect impacts both the static (electronic) and dynamic interprotein electrostatic interactions and significantly lowers the free energy of the barnase-barstar complex.

Understanding the molecular mechanism of enzyme dynamics of ribonuclease A through protonation/deprotonation of HIS48.

Molecular dynamics simulation is carried out to investigate the enzyme dynamics of RNase A with the HIS48 in three different states (HIP48 (protonated), HID48 (deprotonated), and H48A mutant). Insights derived from the current theoretical study, combined with the available experimental observations, enabled us to provide a microscopic picture for the efficient enzyme dynamics. Specifically, in the "closed" state or HIP48, the N-terminal hinge loop is intact and the enzyme remains in a relatively stable conformation which is preferred for catalytic reaction. Deprotonation of HIS48 induces the denaturing of this hinge-loop into a 3(10)-helix, causing it to break the original interaction network around the loop-1 and drive the partial unfolding of the N-terminal. The enhanced dynamic motion of the N-terminal helix facilitates the release of the catalytic product (the rate limiting step) and speeds up the overall catalytic process. The current study established that HIS49 acts as a modulator for the transformation of conformational states through the perturbing of hydrogen bond networks across loop-1, the N-terminal helix, and other residues nearby. Our study suggests that HIS48 may also serve to transport loop-1's kinetic energy to the reaction center.

Electrostatic polarization makes a substantial contribution to the free energy of avidin-biotin binding.

Avidin-biotin is one of the strongest protein-ligand binding systems, with broad applications in biomedical science. Here we report a quantum-based computational study to help elucidate the mechanism of binding avidin to biotin (BTN1) and its close analogue, 2'-iminobiotin (BTN2). Our study reveals that electronic polarization of protein plays a critical role in stabilizing the beta sheet (Thr113-Arg122) at the binding site and makes a substantial contribution to the free energy of avidin-biotin binding. The current finding is in contradiction to the previous notion that electrostatic interaction has no effect on or makes an unfavorable contribution to the free energy of avidin-biotin binding. Our calculations also show that the difference in binding free energy of avidin to BTN1 and BTN2 is almost entirely due to the contribution of electrostatic interaction resulting from polarization-induced stabilization of a hydrogen bond between avidin and BTN1. The current result provides strong evidence that protein polarization accounts for the electrostatic contribution to binding free energy that was missing in previous studies of avidin-biotin binding.

Simulation of NMR data reveals that proteins' local structures are stabilized by electronic polarization.

Molecular dynamics simulations of NMR backbone relaxation order parameters have been carried out to investigate the polarization effect on the protein's local structure and dynamics for five benchmark proteins (bovine pancreatic trypsin inhibitor, immunoglobulin-binding domain (B1) of streptococcal protein G, bovine apo-calbindin D9K, human interleukin-4 R88Q mutant, and hen egg white lysozyme). In order to isolate the polarization effect from other interaction effects, our study employed both the standard AMBER force field (AMBER03) and polarized protein-specific charges (PPCs) in the MD simulations. The simulated order parameters, employing both the standard nonpolarizable and polarized force fields, are directly compared with experimental data. Our results show that residue-specific order parameters at some specific loop and turn regions are significantly underestimated by the MD simulations using the standard AMBER force field, indicating hyperflexibility of these local structures. Detailed analysis of the structures and dynamic motions of individual residues reveals that the hyperflexibility of these local structures is largely related to the breaking or weakening of relevant hydrogen bonds. In contrast, the agreement with the experimental results is significantly improved and more stable local structures are observed in the MD simulations using the polarized force field. The comparison between theory and experiment provides convincing evidence that intraprotein hydrogen bonds in these regions are stabilized by electronic polarization, which is critical to the dynamical stability of these local structures in proteins.

Glycosylation Modulates Human CD2-CD58 Adhesion via Conformational Adjustment.

Human CD2 is a transmembrane cell surface glycoprotein found on T lymphocytes and natural killer cells and plays important roles in immune recognition. The interaction between human CD2 and its counter receptor CD58 facilitates surface adhesion between helper T lymphocytes and antigen presenting cells as well as between cytolytic effectors and target cells. In this study, the molecular effect of glycosylation of CD2 on the structure and dynamics of the CD2-CD58 adhesion complex were examined via MD simulation to help understand the fundamental mechanism of glycosylation that controls CD2-CD58 adhesion. The present result and detailed analysis revealed that the binding interaction of human CD2-CD58 is dominated by three hot spots that form a binding triangle whose topology is critical for stable binding of CD2-CD58. Our study found that the conformation of human CD2, represented by the topology of this binding triangle, is significantly adjusted and steered by glycosylation toward a particular conformation that energetically stabilizes the CD2-CD58 complex. Thus, the fundamental mechanism of glycosylation of human CD2 is to promote CD2-CD58 binding by conformational adjustment of CD2. The current result and explanation are in excellent agreement with previous experiments and help elucidate the dynamical mechanism of glycosylation of human CD2.

Development of an effective polarizable bond method for biomolecular simulation.

An effective polarizable bond (EPB) model has been developed for computer simulation of proteins. In this partial polarizable approach, all polar groups of amino acids are treated as polarizable, and the relevant polarizable parameters were determined by fitting to quantum calculated electrostatic properties of these polar groups. Extensive numerical tests on a diverse set of proteins (including 1IEP, 1MWE, 1NLJ, 4COX, 1PGB, 1K4C, 1MHN, 1UBQ, 1IGD) showed that this EPB model is robust in MD simulation and can correctly describe the structure and dynamics of proteins (both soluble and membrane proteins). Comparison of the computed hydrogen bond properties and dynamics of proteins with experimental data and with results obtained from the nonpolarizable force field clearly demonstrated that EPB can produce results in much better agreement with experiment. The averaged deviation of the simulated backbone N-H order parameter of the B3 immunoglobobulin-binding domain of streptococcal protein G from experimental observation is 0.0811 and 0.0332 for Amber99SB and EPB, respectively. This new model inherited the effective character of the classic force field and the fluctuating feature of previous polarizable models. Different from other polarizable models, the polarization cost energy is implicitly included in the present method. As a result, the present method avoids the problem of over polarization and is numerically stable and efficient for dynamics simulation. Finally, compared to the traditional fixed AMBER charge model, the present method only adds about 5% additional computational time and is therefore highly efficient for practical applications.

Studying the Effect of Site-Specific Hydrophobicity and Polarization on Hydrogen Bond Energy of Protein Using a Polarizable Method.

Quantification of backbone hydrogen bond energies in protein folding has remained elusive despite extensive theoretical and experimental investigations over the past 70 years. This is due to difficulties in experimental mutagenesis study as well as the lack of quantitatively reliable methods in theoretical calculation. Recent advance in experiment has enabled accurate measurement of site-specific backbone hydrogen bond energy in protein. In the present work, we developed an accurate and practical polarizable method to study site-specific hydrogen bond energies in the PIN WW domain. Excellent quantitative agreement between our calculated hydrogen bonding energy and recent experimental measurement is obtained. The direct comparison between theory and experiment helps uncover the microscopic mechanism of experimentally observed context dependent hydrogen bond contribution to protein stability in beta-sheet. In particular, our study reveals two effects that act in a cooperative manner to impact the strength of a hydrogen bond. One is the dynamic stability of the hydrogen bond determined by nearby solvent molecules, and the other is the polarization state of the hydrogen bond influenced by local electrostatic environment. The polar character of the hydrogen bond results in strong coupling between hydrophobic and polarization interactions in a cooperative manner. This nonadditive character in hydrogen bonding should help us better understand the microscopic mechanism in protein folding. Our study also investigated the possible structural effect of backbone amide to ester mutation which should be helpful to experimentalists using this technique in mutagenesis study.

Exploring the molecular mechanism of stabilization of the adhesion domains of human CD2 by N-glycosylation.

N-Glycosylation is one of the most common cotranslational and post-translational modifications occurring in protein biosynthesis and plays a critical role in protein folding and structural diversification. Molecular dynamics studies of two benchmark systems, the NH(2)-terminal human CD2 adhesion domain (HsCD2ad), and the NH(2)-terminal rat CD2 adhesion domain (RnCD2ad) were carried out to investigate the energetic and dynamic effect of N-glycosylation on protein's stability. Our study revealed that N-glycosylation of HsCD2ad at the type I ß-bulge turn strengthens the relevant hydrogen bonds, in particular, the hydrogen bond between Asn(65)OD1-Thr(67)HG1. Dynamic cross correlation map analysis showed that nonglycosylated HsCD2ad has strong anticorrelated motions, whereas glycosylated HsCD2ad largely destroyed this anticorrelated motion. As a result, N-glycosylation energetically and dynamically stabilizes HsCD2ad. In contrast, N-glycosylation of RnCD2ad does not display observable effect on protein's stabilization. The current theoretical result is in excellent agreement with the recent thermodynamic experiment of Culyba et al. and indicates that enthalpy and entropy may both contribute to the stabilization of human CD2 by N-glycosylation.

Quantifying the stabilizing energy of the intraprotein hydrogen bond due to local mutation.

MD simulation of the WW domain of PIN based on a dynamically adjusted polarized protein-specific force field from quantum fragment calculations is carried out in both wild and VAL22ALA mutant states. The result shows that the geometry of the Arg14-TYR23 hydrogen bond is conserved upon mutation of VAL22 to ALA. However, the electrostatic energy of this hydrogen bond in the mutant is found to be 0.6 kcal/mol weaker than in the wild state, in close agreement with the experimentally measured upper limit of 1.2 kcal/mol. Analysis shows that the weakened energy of this hydrogen bond in the mutant is due to its dynamically changed polarization resulting from an altered local electrostatic environment near the hydrogen bond which becomes more exposed to the solvent than in the wild.

NMR scalar coupling constant reveals that intraprotein hydrogen bonds are dynamically stabilized by electronic polarization.

Molecular dynamics simulations based on the standard nonpolarizable AMBER force field and on quantum-derived polarized protein-specific charge (PPC) are performed to compute NMR scalar coupling constants across hydrogen bonds for three benchmark protein systems: ubiquitin, the GB1 domain of protein G, and the SMN Tudor domain. Direct comparison of the simulation result with experimental data gives strong evidence that intraprotein hydrogen bonds are significantly stabilized by electronic polarization, both in terms of NMR scalar coupling constants and X-ray determined geometries of hydrogen bonds. Without the polarization effect in the force field, hydrogen bonds are found to be "too loose", which leads to less stable or even unstable local structures of proteins.

Electronic polarization is important in stabilizing the native structures of proteins.

Quantum mechanical computations of proteins based on the molecular fragment approach have been carried out, and polarized protein-specific charges have been derived to provide accurate electrostatic interactions for a benchmark set of proteins. Our study shows that, under the polarized protein-specific force field, the native structure indeed corresponds to the lowest-energy conformation for these proteins. In contrast, when a standard mean-field force field such as AMBER is used, the energies of many decoy structures of proteins could be lower than those of the native structures. Furthermore, MD simulations were carried out and verified that the native structures of these proteins not only are statically more stable but are also dynamically more stable under the polarized protein-specific force field. The present results, together with several recent studies, provide strong evidence that protein polarization is critical to stabilizing the native structures of proteins.