Ether Hydrolysis, Ether Thiolysis, and the Catalytic Power of Etherases in the Disassembly of Lignin

The recycling of much of the carbon in Nature depends on the breakdown of polymers in woody matter, notably cellulose (a polyacetal) and lignin (a polyether). Here, we show that equilibrium favors ether hydrolysis in water, although the rates of spontaneous hydrolysis of ethers are too slow to measure in neutral solution except at temperatures approaching the critical point of water. Circumventing that kinetic obstacle, glutathione-dependent etherases from white-rot fungi are known to employ the thiolate group of glutathione to attack guaiacyl ethers. Experiments at elevated temperatures indicate that thioglycolate attacks diethyl ether in water, in the absence of enzymes, with a rate constant of 6 × 10^-11 M^-1 s^-1 at 25 °C and that ether thiolysis is strongly favored thermodynamically, with a K_eq value of 2.5 × 10⁶ (ΔG = -8.7 kcal/mol). Compared with the rate of non-enzymatic thiolysis, the lignin-degrading etherases LigE and LigF produce 10¹⁵-fold rate enhancements, among the largest that have been observed for an enzyme acting on two substrates.