Dynamics Of HIV-1 Protease Research Articles

Effects of drug-resistant mutations on the dynamic properties of HIV-1 protease and inhibition by Amprenavir and Darunavir.

Molecular dynamics simulations are performed to investigate the dynamic properties of wild-type HIV-1 protease and its two multi-drug-resistant variants (Flap + (L10I/G48V/I54V/V82A) and Act (V82T/I84V)) as well as their binding with APV and DRV inhibitors. The hydrophobic interactions between flap and 80 s (80’s) loop residues (mainly I50-I84’ and I50’-I84) play an important role in maintaining the closed conformation of HIV-1 protease. The double mutation in Act variant weakens the hydrophobic interactions, leading to the transition from closed to semi-open conformation of apo Act. APV or DRV binds with HIV-1 protease via both hydrophobic and hydrogen bonding interactions. The hydrophobic interactions from the inhibitor is aimed to the residues of I50 (I50’), I84 (I84’), and V82 (V82’) which create hydrophobic core clusters to further stabilize the closed conformation of flaps, and the hydrogen bonding interactions are mainly focused with the active site of HIV-1 protease. The combined change in the two kinds of protease-inhibitor interactions is correlated with the observed resistance mutations. The present study sheds light on the microscopic mechanism underlying the mutation effects on the dynamics of HIV-1 protease and the inhibition by APV and DRV, providing useful information to the design of more potent and effective HIV-1 protease inhibitors.

Interchain hydrophobic clustering promotes rigidity in HIV-1 protease flap dynamics: new insights from Molecular Dynamics

The dynamics of HIV-1 protease (HIV-pr), a drug target for HIV infection, has been studied extensively by both computational and experimental methods. The flap dynamics of HIV-pr is considered to be more important for better ligand binding and enzymatic actions. Moreover, it has been demonstrated that the drug-induced mutations can change the flap dynamics of HIV-pr affecting the binding affinity of the ligands. Therefore, detailed understanding of flap dynamics is essential for designing better inhibitors. Previous computational investigations observed significant variation in the flap opening in nanosecond time scale indicating that the dynamics is highly sensitive to the simulation protocols. To understand the sensitivity of the flap dynamics on the force field and simulation protocol, molecular dynamics simulations of HIV-pr have been performed with two different AMBER force fields, ff99 and ff02. Two different trajectories (20 ns each) were obtained using the ff99 and ff02 force field. The results showed polarizable force field (ff02) make the flap tighter than the nonpolarizable force field (ff99). Some polar interactions and hydrogen bonds involving flap residues were found to be stronger with ff02 force field. The formation of interchain hydrophobic cluster (between flap tip of one chain and active site wall of another chain) was found to be dominant in the semi-open structures obtained from the simulations irrespective of the force field. It is proposed that an inhibitor, which will promote this interchain hydrophobic clustering, may make the flaps more rigid, and presumably the effect of mutation would be small on ligand binding.

CONFORMATIONAL DYNAMICS OF HIV-1 PROTEASE: A COMPARATIVE MOLECULAR DYNAMICS SIMULATION STUDY WITH MULTIPLE AMBER FORCE FIELDS

Flap dynamics of HIV-1 protease (HIV-pr) controls the entry of inhibitors and substrates to the active site. Dynamical models from previous simulations are not all consistent with each other and not all are supported by the NMR results. In the present work, the effect of force field on the dynamics of HIV-pr is investigated by MD simulations using three AMBER force fields ff99, ff99SB, and ff03. The generalized order parameters for amide backbone are calculated from the three force fields and compared with the NMR S2 values. We found that the ff99SB and ff03 force field calculated order parameters agree reasonably well with the NMR S2 values, whereas ff99 calculated values deviate most from the NMR order parameters. Stereochemical geometry of protein models from each force field also agrees well with the remarks from NMR S2 values. However, between ff99SB and ff03, there are several differences, most notably in the loop regions. It is found that these loops are, in general, more flexible in the ff03 force field. This results in a larger active site cavity in the simulation with the ff03 force field. The effect of this difference in computer-aided drug design against flexible receptors is discussed.

Backbone 1H, 13C, and 15N chemical shift assignment for HIV-1 protease subtypes and multi-drug resistant variant MDR 769

HIV-1 protease (HIV-1PR) is an essential drug target in the treatment of patients infected with HIV-1. Mutations are found to arise in over 38 of 99 amino acid sites in this protein in response to drug therapy or natural selection, where many are found combinations that alter enzyme kinetics or inhibitor susceptibility without a clear structural mechanism. In efforts to understand how these mutations alter the flexibility and dynamics of HIV-1PR, we report the backbone (1)H, (13)C, and (15)N chemical shift assignments for subtypes C, circulating recombinant form CRF01_AE and a multi-drug resistant variant MDR 769. These assignments are essential for future work aimed at characterizing backbone dynamics, exchange dynamics and dynamics of protein/substrate or protein/inhibitor interactions.

Insights into the Dynamics of HIV-1 Protease: A Kinetic Network Model Constructed from Atomistic Simulations

The conformational dynamics in the flaps of HIV-1 protease plays a crucial role in the mechanism of substrate binding. We develop a kinetic network model, constructed from detailed atomistic simulations, to determine the kinetic mechanisms of the conformational transitions in HIV-1 PR. To overcome the time scale limitation of conventional molecular dynamics (MD) simulations, our method combines replica exchange MD with transition path theory (TPT) to study the diversity and temperature dependence of the pathways connecting functionally important states of the protease. At low temperatures the large-scale flap opening is dominated by a small number of paths; at elevated temperatures the transition occurs through many structurally heterogeneous routes. The expanded conformation in the crystal structure 1TW7 is found to closely mimic a key intermediate in the flap-opening pathways at low temperature. We investigated the different transition mechanisms between the semi-open and closed forms. The calculated relaxation times reveal fast semi-open ↔ closed transitions, and infrequently the flaps fully open. The ligand binding rate predicted from this kinetic model increases by 38-fold from 285 to 309 K, which is in general agreement with experiments. To our knowledge, this is the first application of a network model constructed from atomistic simulations together with TPT to analyze conformational changes between different functional states of a natively folded protein.

COARSE GRAINED MODELING OF BIOPOLYMERS AND PROTEINS: METHODS AND APPLICATIONS

Single-molecule mechanics of DNA/RNA, polypeptide and protein has become a new frontier in life sciences. There is an increasing need in participant of researcher in applied mechanics to perform a systematic study on methodologies and basic sciences in molecular mechanics. The wide-applied continuum mechanics may not be applicable in study of the biomolecules. At the same time, the all-atom molecular dynamics method is severely limited by the size and time scales of systems it can simulate. As an alternative, coarse-grained models have recently enjoyed intense interest. Coarse-grained representations combined with enhanced computer power currently allow the simulation of systems of biologically relevant size (submicrometric) and timescale (microsecond or millisecond). In this paper, we reported some progresses we made in the modeling of biomolecules using the coarse-grained methods including, the collapse of biopolymers, the dynamics of HIV-1 protease and the binding dynamics of inhibitors into the protease, aiming to stimulating a broader interest in developing molecular mechanics.

Flap opening mechanism of HIV-1 protease

The active site of aspartic proteases, such as HIV-1 protease (PR), is covered by one or more flaps, which restrict access to the active site. For HIV-1 PR, X-ray diffraction studies suggested that in the free enzyme the two flaps are packed onto each other loosely in a semi-open conformation, while molecular dynamics (MD) studies observed that the flaps can also separate into open conformations. In this study, the mechanism of flap opening and the structure and dynamics of HIV-1 PR with semi-open and open flap conformations were investigated using molecular dynamics simulations. The flaps showed complex dynamic behavior as two distinct mechanisms of flap opening and various stable flap conformations (semi-open, open and curled) were observed during the simulations. A network of weakly polar interactions between the flaps were proposed to be responsible for stabilizing the semi-open flap conformation. It is hypothesized that such interactions could be responsible for making flap opening a highly sensitive gating mechanism which control access to the active site.

A molecular dynamics study of the structural stability of HIV‐1 protease under physiological conditions: The role of Na+ ions in stabilizing the active site

HIV-1 protease is most active under weakly acidic conditions (pH 3.5-6.5), when the catalytic Asp25 and Asp25' residues share 1 proton. At neutral pH, this proton is lost and the stability of the structure is reduced. Here we present an investigation of the effect of pH on the dynamics of HIV-1 protease using MD simulation techniques. MD simulations of the solvated HIV-1 protease with the Asp25/25' residues monoprotonated and deprotonated have been performed. In addition we investigated the effect of the inclusion of Na(+) and Cl(-) ions to mimic physiological salt conditions. The simulations of the monoprotonated form and deprotonated form including Na(+) show very similar behavior. In both cases the protein remained stable in the compact, "self-blocked" conformation in which the active site is blocked by the tips of the flaps. In the deprotonated system a Na(+) ion binds tightly to the catalytic dyad shielding the repulsion between the COO(-) groups. Ab initio calculations also suggest the geometry of the active site with the Na(+) bound closely resembles that of the monoprotonated case. In the simulations of the deprotonated form (without Na(+) ions), a water molecule bound between the Asp25 Asp25' side-chains. This disrupted the dimerization interface and eventually led to a fully open conformation.

Cooperative fluctuations of unliganded and substrate-bound HIV-1 protease: a structure-based analysis on a variety of conformations from crystallography and molecular dynamics simulations.

The dynamics of HIV-1 protease, both in unliganded and substrate-bound forms have been analyzed by using an analytical method, Gaussian network model (GNM). The method is applied to different conformations accessible to the protein backbone in the native state, observed in crystal structures and snapshots from fully atomistic molecular dynamics (MD) simulation trajectories. The modes of motion obtained from GNM on different conformations of HIV-1 protease are conserved throughout the MD simulations. The flaps and 40's loop of the unliganded HIV-1 protease structure are identified as the most mobile regions. However, in the liganded structure these flaps lose mobility, and terminal regions of the monomers become more flexible. Analysis of the fast modes shows that residues important for stability are in the same regions of all the structures examined. Among these, Gly86 appears to be a key residue for stability. The contribution of residues in the active site region and flaps to the stability is more pronounced in the substrate-bound form than in the unliganded form. The convergence of modes in GNM to similar regions of HIV-1 protease, regardless of the conformation of the protein, supports the robustness of GNM as a potentially useful and predictive tool.

P-A3-09 Influence of inhibitor binding on HIV-1 protease dynamics: Ringhofer, S. 1, Dutzler, R. 1, Kallen, J. 1, Andrew, P. 3, Visser, A. 4, Billich, A. 3, Scholz, D. 3, Gstach, H. 3, Steinhauser, O. 1, Schreiber, H. 1, Aucr, M. 3, Kungl, A. 3 1 Inst. Theoret. Chem., Univ. Vienna (A); 2 Sandoz Pharma, Basel (CH); 3 Sandoz Forschungs-Inst.,

P-A3-09 Influence of inhibitor binding on HIV-1 protease dynamics: Ringhofer, S. 1, Dutzler, R. 1, Kallen, J. 1, Andrew, P. 3, Visser, A. 4, Billich, A. 3, Scholz, D. 3, Gstach, H. 3, Steinhauser, O. 1, Schreiber, H. 1, Aucr, M. 3, Kungl, A. 3 1 Inst. Theoret. Chem., Univ. Vienna (A); 2 Sandoz Pharma, Basel (CH); 3 Sandoz Forschungs-Inst.,