Thiol and amino analogs as alternate substrates for glycerokinase from Candida mycoderma

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Abstract
The kinetic and catalytic mechanism of glycerokinase from Candida mycoderma was examined with thiol and amino analogues of glycerol and with MgAMPPCP, an analogue of MgATP. (S)-1- Aminopropanediol was phosphorylated on nitrogen (Vmax 0.4% that of glycerol) while the R enantiomer was phosphorylated on oxygen (Vmax 0.7% that of glycerol). (S)-1-Mercaptopropanediol was phosphorylated on oxygen (Vmax 3.5% that of glycerol), while the R enantiomer was phosphorylated on sulfur (Vmax 0.001% that of glycerol). The hydroxyl group at C-2 thus orients the substrate in the active site, while that at the carbon remote phosphorylation enhances both catalysis and binding of the substrate, presumably because of hydrogen-bonding interactions. The kinetic mechanism is random with a high degree of synergistic binding between the substrates, so that the mechanism appears ordered with glycerol adding first but equilibrium ordered with MgATP binding first with the amino analogues.