Mechanism of the myosin catalyzed hydrolysis of ATP as rationalized by molecular modeling

Abstract

The intrinsic chemical reaction of adenosine triphosphate (ATP) hydrolysis catalyzed by myosin is modeled by using a combined quantum mechanics and molecular mechanics (QM/MM) methodology that achieves a near ab initio representation of the entire model. Starting with coordinates derived from the heavy atoms of the crystal structure (Protein Data Bank ID code 1VOM) in which myosin is bound to the ATP analog ADP⅐VO 4 ؊ , a minimum-energy path is found for the transformation ATP ؉ H 2O 3 ADP ؉ Pi that is characterized by two distinct events: (i) a low activation-energy cleavage of the P ␥OO␤␥ bond and separation of the ␥-phosphate from ADP and (ii) the formation of the inorganic phosphate as a consequence of proton transfers mediated by two water molecules and assisted by the Glu-459 -Arg-238 salt bridge of the protein. The minimum-energy model of the enzyme-substrate complex features a stable hydrogen-bonding network in which the lytic water is positioned favorably for a nucleophilic attack of the ATP ␥-phosphate and for the transfer of a proton to stably bound second water. In addition, the P␥OO␤␥ bond has become significantly longer than in the unbound state of the ATP and thus is predisposed to cleavage. The modeled transformation is viewed as the part of the overall hydrolysis reaction occurring in the closed enzyme pocket after ATP is bound tightly to myosin and before conformational changes preceding release of inorganic phosphate.ATP hydrolysis ͉ enzymatic catalysis ͉ energy profile ͉ quantum mechanics and molecular mechanics simulations T he mechanism of hydrolysis of adenosine triphosphate (ATP) by myosin, leading to adenosine diphosphate (ADP) and inorganic phosphate (P i ), which constitutes one of the most important enzymatic reactions responsible for energy transduction into the directed movements of adjoining actin filaments, continues to remain a subject of active debates (1-16), a significant part of which relates to what constitutes the acceptor of the proton that must be released by the ''lytic'' water in its nucleophilic attack on the ATP ␥-phosphate.In terms of the generally accepted kinetic scheme (1-3), the relevant ATP-myosin transformations may be described by the equationin which M* and M** indicate conformers of myosin. As reported (1-3), reaction (Eq. 1) occurs with a near unit equilibrium constant K Ͻ 10 and the estimated rate constants k ϩ Ն 160 s Ϫ1 and k Ϫ Ն 18 s Ϫ1 . The rate constant k ϩ ϭ 160 s Ϫ1 can be converted to the free-energy activation barrier ⌬G # Ϸ 14.6 kcal/mol at room temperature, T ϭ 300 K, by applying a simple transition-state theory formula (17) k Ϸ 6⅐10 12 exp͓Ϫ⌬G # ͞RT͔. [2]However, noting that the experimental rate constants of reaction (Eq. 1) incorporate contributions from conformational changes in the protein from M* to M** leads us to expect that the activation energy of the intrinsic chemical reactionwhich excludes conformational rearrangements, should be considerably Ͻ14.6 kcal/mol. However, previous attempts (13, 15, 16) to simulate the mechanism of react...

Why nature really chose phosphate

“…25 is drastically lowered and the second step becomes rate limiting). This problem become even more apparent in a study of ATP hydrolysis (Grigorenko et al 2007b), (discussed in Section 4.2) and in the EF-Tu study discussed below. Incidentally, the solution study that produced a barrier of 20 kcal mol x1 (Grigorenko et al 2006) instead of about 28 kcal mol x1 was done by energy minimization, which is unacceptable for the enormous dimensionality of the cluster used, and, in fact, assuming that the proposed mechanism of PT along a chain of three water molecules in water can be explored by energy minimization in somewhat unrealistic.…”

Mutations in Sarcomere Protein Genes in Left Ventricular Noncompaction

“…Absence of Asp239 could change the steric structure of the binding site and interfere with ATP binding. The positively charged residue Arg243 is involved in phosphate binding, 22 and ATP binding may be diminished if the residue is changed for a histidine. Phe252 is located in a loop that is part of a hydrophobic cluster at the outer surface of the ATP binding cleft.…”

Catalytic strategy used by the myosin motor to hydrolyze ATP

“…2B). The simulations have shown that only dissociative pathways have transition barriers that are low enough to be consistent with the experimental rates (9,18,19). Recently, we have explained how myosin achieves the stabilization of the metaphosphate (P γ O 3 − ) that is generated by the dissociative mechanism (20).…”

“…Understanding how myosin achieves its ATPase function is necessary to understand how myosin works as a motor, but also helps one to understand the functioning of the multitude of other nucleotide hydrolyzing enzymes. The catalytic mechanism of ATP hydrolysis in myosin has been studied extensively with methods such as protein crystallography (11)(12)(13)(14), mutagenesis (15, 16), photochemical kinetics (10), and quantum-mechanical simulations of the active site (9,(17)(18)(19)(20)(21). This has yielded structures of analogs of the reactant and transition states, a list of residues affecting the catalytic process, and a number of proposed catalytic pathways.…”

“…Whereas Glu459 has been proposed to act as a general base in the catalysis, the exact role played by the other residues remained unclear. Both associative (9,17,21) and dissociative (18,19) pathways in myosin have been investigated by combined quantum-classical (QM/MM) simulations. In an associative mechanism, the breaking of the P β -O βγ bond is concerted with the attack of water onto the γ-phosphate ( Fig.…”