Mechanism of product specificity of AdoMet methylation catalyzed by lysine methyltransferases: Transcriptional factor p53 methylation by histone lysine methyltransferase SET7/9

摘要

The catalysis by SET7/9 histone lysine methyltransferase of AdoMet N-methylation of the transcriptional factor p53-Lys4-NH2 has been investigated with particular attention paid to the means of product specificity. After formation of the SET7/9 center dot p53-Lys4-NH3(+)center dot AdoMet complex, the following events occur: (i) the appearance of a water channel, (ii) a depronation of p53-Lys4-NH3+ via this water channel into the aqueous solvent, and (iii) AdoMet methylation of p53-Lys4-NH2) to form p53-Lys4-N(Me)H-2(+). The formation of a water channel does not occur on formation of the SET7/9 center dot p53-Lys4-NH3+, SET7/9 center dot p53-Lys4-N(Me)H-2(+)center dot AdoHcy, or SET7/9 center dot p53-Lys4-N(Me)H-2(+)center dot AdoMet complex. Without a water channel, the substrate p53-Lys4-N(Me)H is not available because the proton dissociation p53-Lys4-N(Me)H-2(+)-> p53-Lys4-N(Me)H + H+ does not occur. The lack of formation of a water channel is due to the positioning of the methyl substituent of the SET7/9 center dot p53-Lys4-N(Me)H-2(+)center dot AdoMet complex. By quantum mechanics/molecular mechanics, the computed free energy barrier of the methyl transfer reaction [p53-Lys4-NH2 + AdoMet -> p53-Lys4-N(Me)H-2(+) + AdoHcy] in the SET7/9 complex is Delta G(double dagger) = 20.1 +/- 2.9 kcal/mol. This Delta G(double dagger) is in agreement with the value of 20.9 kcal/mol calculated from the experimental rate constant (1.2 +/- 0.1 min(-1)). Our bond-order computations establish that the methyl transfer reaction in protein lysine methyltransferases occurs via a linear S(N)2 associative reaction with bond making of similar to 50%.