Bearne Stephen L, Spiteri Raymond J
Department of Biochemistry and Molecular Biology, Dalhousie University, Halifax, Nova Scotia, Canada B3H 4H7.
J Theor Biol. 2005 Apr 21;233(4):563-71. doi: 10.1016/j.jtbi.2004.11.003. Epub 2004 Dec 25.
Many enzymes catalyse the heterolytic abstraction of the alpha-proton from a carbon acid substrate. Gerlt and Gassman have applied Marcus formalism to such proton transfer reactions to argue that transition states for concerted general acid-general base catalysed enolization at enzyme active sites occur late on the reaction coordinate (J. Am. Chem. Soc. 115 (1993) 11552). We postulate that as an enzyme evolves, it may decrease deltaG++ for a proton transfer step associated with substrate enolization by following the path of steepest descent on the two-dimensional surface corresponding to deltaG++, as defined by Marcus formalism. We show that for an enzyme that has decreased deltaG++ following the path of steepest descent, the values of the intrinsic kinetic (deltaG++(int,E)) and thermodynamic (deltaG(E)0) barriers for proton transfer reactions on the enzyme may be predicted from the known values of deltaG++(int,N) and deltaG(N)0 for the corresponding non-enzymic reaction and the free energy of activation on the enzyme (deltaG++(E)). In addition, the enzymic transition state will occur later on the reaction coordinate than the corresponding non-enzymic transition state (i.e. x++(E)>x++(N)) if the condition (6 - square root 2)/8<x++(N)<(6 + square root 2)/8 is satisfied. For enzyme-catalysed abstraction of the alpha-proton from carbon acid substrates with high pK(a) values (e.g. pK(a) approximately 29), the free energy of activation for the non-enzymic reaction (deltaG++(N)) is dominated by deltaG(N)0. Reduction of deltaG++, via the path of steepest descent will reduce deltaG0 to a greater extent (i.e. differential binding) than deltaG++(int) if deltaG(N)0>2deltaG++(int,N).
许多酶催化从碳酸底物上异裂提取α-质子。格尔特和加斯曼已将马库斯形式主义应用于此类质子转移反应，以论证在酶活性位点协同一般酸-一般碱催化烯醇化的过渡态在反应坐标上出现较晚（《美国化学会志》115 (1993) 11552）。我们推测，随着酶的进化，它可能通过在由马库斯形式主义定义的对应于ΔG‡的二维表面上沿最陡下降路径，来降低与底物烯醇化相关的质子转移步骤的ΔG‡。我们表明，对于沿最陡下降路径降低了ΔG‡的酶，酶上质子转移反应的本征动力学（ΔG‡(int,E)）和热力学（ΔG(E)0）能垒值可根据相应非酶反应的ΔG‡(int,N)和ΔG(N)0的已知值以及酶上的活化自由能（ΔG‡(E)）来预测。此外，如果满足条件(6 - √2)/8 < x‡(N) < (6 + √2)/8，则酶催化的过渡态在反应坐标上比相应的非酶催化过渡态出现得更晚（即x‡(E) > x‡(N)）。对于从具有高pK(a)值（例如pK(a)约为29）的碳酸底物上酶催化提取α-质子，非酶反应的活化自由能（ΔG‡(N)）主要由ΔG(N)0决定。通过最陡下降路径降低ΔG‡，如果ΔG(N)0 > 2ΔG‡(int,N)，将比ΔG‡(int)更大程度地降低ΔG0（即差异结合）。
