Free Energy Simulations of a GTPase: GTP and GDP Binding to Archaeal Initiation Factor 2

International audienceArchaeal initiation factor 2 (aIF2) is a protein involved in the initiation of protein biosynthesis. In its GTP-bound, "ON" conformation, aIF2 binds an initiator tRNA and carries it to the ribosome. In its GDP-bound, "OFF" conformation, it dissociates from tRNA. To understand the specific binding of GTP and GDP and its dependence on the ON or OFF conformational state of aIF2, molecular dynamics free energy simulations (MDFE) are a tool of choice. However, the validity of the computed free energies depends on the simulation model, including the force field and the boundary conditions, and on the extent of conformational sampling in the simulations. aIF2 and other GTPases present specific difficulties; in particular, the nucleotide ligand coordinates a divalent Mg(2+) ion, which can polarize the electronic distribution of its environment. Thus, a force field with an explicit treatment of electronic polarizability could be necessary, rather than a simpler, fixed charge force field. Here, we begin by comparing a fixed charge force field to quantum chemical calculations and experiment for Mg(2+):phosphate binding in solution, with the force field giving large errors. Next, we consider GTP and GDP bound to aIF2 and we compare two fixed charge force fields to the recent, polarizable, AMOEBA force field, extended here in a simple, approximate manner to include GTP. We focus on a quantity that approximates the free energy to change GTP into GDP. Despite the errors seen for Mg(2+):phosphate binding in solution, we observe a substantial cancellation of errors when we compare the free energy change in the protein to that in solution, or when we compare the protein ON and OFF states. Finally, we have used the fixed charge force field to perform MDFE simulations and alchemically transform GTP into GDP in the protein and in solution. With a total of about 200 ns of molecular dynamics, we obtain good convergence and a reasonable statistical uncertainty, comparable to the force field uncertainty, and somewhat lower than the predicted GTP/GDP binding free energy differences. The sign and magnitudes of the differences can thus be interpreted at a semiquantitative level, and are found to be consistent with the experimental binding preferences of ON- and OFF-aIF2

Possibility of proton passage through all metal aromatic Al 42- ring in HAl4-

International audienceWe report theoretical investigations on HAl 4- in which the proton can move through the Al 42- ring similar to umbrella inversion. The potential energy for the motion is a double well, with an activation barrier of 4.45 kcal/mol. We find that excitation to v = 6 vibrational levels should lead to easy passage of H+ through the ring. After considering the tunnelling effect, inversion rate at 298 K is calculated using transition state theory and is found to be 1.3 × 1010/s-1. © 2010 Wiley Periodicals, Inc

Dcab(2-) - Substituted manganese vinylidene complexes as better electron reservoirs: A DFT study

Mononuclear vinylidene complexes of the type Mn(C5H4R')(R"2PCH2CH2P R"(2))= C = C(R')(H) are synthesized and reported by Venkatesan et a]. as potential electron reservoirs, which can store and release electrons in a reversible fashion. However, the slow oxidation of the parent compound leads to an undesired product. In our model compound Mn(C5H5)(PH3)(2 =)C = C(R)(H), we substituted the cyclopentadienyl moiety by the isolobal dianionic dicarbollyl ligand Dcab(2-) (C2B9H112-) and investigated whether this simple substitution can reduce the production of an undesired product. Our calculations of vertical electron detachment energy, thermodynamic feasibility, and molecular orbital analysis (with substituents R = H, Me, Ph on the Cp atom of our model system) show that the substitution is thermodynamically favorable and leads to easy oxidation of the parent compound, easy dimerization, and better reversibility. Our comparative study between Mn(C5H5)(PH3)(2 =)C = C(R)(H) and Mn(Dcab)(PH3)(2) = C = C(R)(H)(-) (where R = H. Me, Ph) predicts the latter to be a better electronic reservoir

Structure and bonding of MCB5H7MCB_5H_7 and its sandwiched dimer CB5H6M−MCB5H6CB_5H_6M-MCB_5H_6 (M = Si, Ge, Sn): Isomer stability and preference for slip distorted structure

We find sandwiched metal dimers $CB_5H_6M-MCB_5H_6$ (M = Si, Ge, Sn) which are minima in the potential energy surface with a characteristic M–M single bond. The NBO analysis and the M–M distances (\AA) (2.3, 2.44 and 2.81 for M = Si, Ge, Sn) indicate substantial M–M bonding. Formal generation of $CB_5H_6M-MCB_5H_6$ has been studied theoretically. Consecutive substitution of two boron atoms in $B_7H^{-2}_7$ by M (Si, Ge, Sn) and carbon, respectively followed by dehydrogenation may lead to our desired $CB_5H_6M- MCB_5H_6$. We find that the slip distorted geometry is preferred for $MCB_5H_7$ and its dehydrogenated dimer $CB_5H_6M -MCB_5H_6$. The slip-distortion of M-M bond in $CB_5H_6M-MCB_5H_6$ is more than the slip distortion of M–H bond in MCB5H7. Molecular orbital analysis has been done to understand the slip distortion. Larger M–M bending ($CB_5H_6M -MCB_5H_6$) in comparison with M–H bending ($MCB_5H_7$) is suspected to be encouraged by stabilization of one of the M–M π bonding MO’s. Preference of M to occupy the apex of pentagonal skeleton of $MCB_5H_7$ over its icosahedral analogue $MCB_{10}H_{11}$ has been observed

Why base tautomerization does not cause errors in mRNA decoding on the ribosome

The structure of the genetic code implies strict Watson-Crick base pairing in the first two codon positions, while the third position is known to be degenerate, thus allowing wobble base pairing. Recent crystal structures of near-cognate tRNAs accommodated into the ribosomal A-site, however, show canonical geometry even with first and second position mismatches. This immediately raises the question of whether these structures correspond to tautomerization of the base pairs. Further, if unusual tautomers are indeed trapped why do they not cause errors in decoding? Here, we use molecular dynamics free energy calculations of ribosomal complexes with cognate and near-cognate tRNAs to analyze the structures and energetics of G-U mismatches in the first two codon positions. We find that the enol tautomer of G is almost isoenergetic with the corresponding ketone in the first position, while it is actually more stable in the second position. Tautomerization of U, on the other hand is highly penalized. The presence of the unusual enol form of G thus explains the crystallographic observations. However, the calculations also show that this tautomer does not cause high codon reading error frequencies, as the resulting tRNA binding free energies are significantly higher than for the cognate complex

Nucleotide recognition by the initiation factor aIF5B: Free energy simulations of a neoclassical GTPase.

International audienceThe GTPase aIF5B is a universally conserved initiation factor that assists ribosome assembly. Crystal structures of its nucleotide complexes, X-ray(GTP) and X-ray(GDP), are similar in the nucleotide vicinity, but differ in the orientation of a distant domain IV. This has led to two, contradictory, mechanistic models. One postulates that X-ray(GTP) and X-ray(GDP) are, respectively, the active, "ON" and the inactive, "OFF" states; the other postulates that both structures are OFF, whereas the ON state is still uncharacterized. We study GTP/GDP binding using molecular dynamics and a continuum electrostatic free energy method. We predict that X-ray(GTP) has a ≈ 3 kcal/mol preference to bind GDP, apparently contradicting its assignment as ON. However, the preference arises mainly from a single, nearby residue from the switch 2 motif: Glu81, which becomes protonated upon GTP binding, with a free energy cost of about 4 kcal/mol. We then propose a different model, where Glu81 protonation/deprotonation defines the ON/OFF states. With this model, the X-ray(GTP):GTP complex, with its protonated Glu81, is ON, whereas X-ray(GTP):GDP is OFF. The model postulates that distant conformational changes such as domain IV rotation are "uncoupled" from GTP/GDP exchange and do not affect the relative GTP/GDP binding affinities. We analyze the model using a general thermodynamic framework for GTPases. It yields rather precise predictions for the nucleotide specificities of each state, and the state specificities of each nucleotide, which are roughly comparable to the homologues IF2 and aIF2, despite the lack of any conformational switching in the model. © 2012 Wiley Periodicals, Inc

Stabilization of an All-Metal Antiaromatic Molecule (Al4Li4) Using BH and C as Caps

It has been reported by Pati et al. (J. Am. Chem. Soc. 2005, 127, 3496) that coordination with a transition metal can stabilize the “antiaromatic”, all-metal compound Al4Li4. Here, we report that it can also be stabilized by capping with a main group element like C and its isoelectronic species BH. Our calculations of binding energy, nuclear independent chemical shift, energy decomposition analysis, and molecular orbital analysis support the capping-induced stability, reduction of bond length alternation, and increase of aromaticity of these BH/C-capped Al4Li4 systems. The interaction between px and py orbitals of BH/C and the HOMO and LUMO of Al4Li4 is responsible for the stabilization. Our calculations suggest that capping can introduce fluxionality at room temperature

Conformational selection through electrostatics: Free energy simulations of GTP and GDP binding to archaeal initiation factor 2.

International audienceArchaeal Initiation Factor 2 is a GTPase involved in protein biosynthesis. In its GTP-bound, "ON" conformation, it binds an initiator tRNA and carries it to the ribosome. In its GDP-bound, "OFF" conformation, it dissociates from tRNA. To understand the specific binding of GTP and GDP and their dependence on the conformational state, molecular dynamics free energy simulations were performed. The ON state specificity was predicted to be weak, with a GTP/GDP binding free energy difference of -1 kcal/mol, favoring GTP. The OFF state specificity is larger, 4 kcal/mol, favoring GDP. The overall effects result from a competition among many interactions in several complexes. To interpret them, we use a simpler, dielectric continuum model. Several effects are robust with respect to the model details. Both nucleotides have a net negative charge, so that removing them from solvent into the binding pocket carries a desolvation penalty, which is large for the ON state, and strongly disfavors GTP binding compared to GDP. Short-range interactions between the additional GTP phosphate group and ionized sidechains in the binding pocket offset most, but not all of the desolvation penalty; more distant groups also contribute significantly, and the switch 1 loop only slightly. The desolvation penalty is lower for the more open, wetter OFF state, and the GTP/GDP difference much smaller. Short-range interactions in the binding pocket and with more distant groups again make a significant contribution. Overall, the simulations help explain how conformational selection is achieved with a single phosphate group

Tunneling of three borons in a B12B_{12} cluster

We report a theoretical investigation on a $B_{12}$ cluster, which can exhibit a through ring umbrella inversion. Our calculations show that a part of the molecule consisting of a three membered boron ring can invert through the rest, viz., a nine membered boron ring. Using a simple model we calculate the double well potential for the motion. The barrier for inversion is found to be 4.31 kcal/mol. The vibrational levels and tunneling splitting are calculated using this potential. We find that the vibrational excitation to the v = 17 level will cause large amplitude “inversion oscillation” of the molecule. After including the tunneling effect, inversion rate at 298 K is calculated using transition state theory and is found to be $1.17 X 10^{10}/s$

Tunneling of three borons in a B<SUB>12</SUB> cluster

We report a theoretical investigation on a B12 cluster, which can exhibit a through ring umbrella inversion. Our calculations show that a part of the molecule consisting of a three membered boron ring can invert through the rest, viz., a nine membered boron ring. Using a simple model we calculate the double well potential for the motion. The barrier for inversion is found to be 4.31 kcal/mol. The vibrational levels and tunneling splitting are calculated using this potential. We find that the vibrational excitation to the v = 17 level will cause large amplitude "inversion oscillation" of the molecule. After including the tunneling effect, inversion rate at 298 K is calculated using transition state theory and is found to be 1.17 &#215; 1010/s