Catalytic mechanism and product specificity of rubisco large subunit methyltransferase: QM/MM and MD investigations

Molecular dynamics (MD) simulations and hybrid quantum mechanics/molecular mechanics (QM/MM) calculations have been carried out in an investigation of Rubisco large subunit methyltransferase (LSMT). It was found that the appearance of a water channel is required for the stepwise methylation by S-adenosylmethionine (AdoMet). The water channel appears in the presence of AdoMet (LSMT.Lys-NH3+.AdoMet), but is not present immediately after methyl transfer (LSMT.Lys-N(Me)H2+.AdoHcy). The water channel allows proton dissociation from both LSMT.AdoMet.Lys-NH3+ and LSMT.AdoMet.Lys-N(Me)H2+. The water channel does not appear for proton dissociation from LSMT.AdoMet.Lys-N(Me)2H+, and a third methyl transfer does not occur. By QM/MM, the calculated free energy barrier of the first methyl transfer reaction catalyzed by LSMT (Lys-NH2 + AdoMet --> Lys-N(Me)H2+ + AdoHcy) is DeltaG++ = 22.8 +/- 3.3 kcal/mol. This DeltaG++ is in remarkable agreement with the value 23.0 kcal/mol calculated from the experimental rate constant (6.2 x 10-5 s-1). The calculated DeltaG++ of the second methyl transfer reaction (AdoMet + Lys-N(Me)H --> AdoHcy + Lys-N(Me)2H+) at the QM/MM level is 20.5 +/- 3.6 kcal/mol, which is in agreement with the value 22.0 kcal/mol calculated from the experimental rate constant (2.5 x 10-4 s-1). The third methyl transfer (Lys-N(Me)2 + AdoMet --> Lys-N(Me)3+ + AdoHcy) is associated with an allowed DeltaG++ of 25.9 +/- 3.2 kcal/mol. However, this reaction does not occur because a water channel does not form to allow the proton dissociation of Lys-N(Me)2H+. Future studies will determine whether the product specificity of lysine (mono, di, and tri) methyltransferases is determined by the formation of water channels.