THE MECHANISMS OF HYDROLYSIS OF GLYCERALDEHYDE-3-PHOSPHATE

The pH rate profile of the unbuffered hydrolysis of glyceraldehyde-3-p hosphate (G3P) has been studied at 49.4 degrees C in the pH range 0-11 . The form of the profile at pH<4 is typical of hydrolyses of the mono anion of phosphate monoesters. Similarly as other sugar phosphates, at pH 7-8 there is a spontaneous reaction of the phosphate dianion, wher e the only product is glyceraldehyde, followed at higher pH by hydroxi de-catalyzed hydrolysis that produces methylglyoxal. Products have bee n analyzed by paper chromatography, GLC, periodate oxidation, and a mo dified reaction with <<Purpald>>. No solvent isotope effect (SIE) has been observed for the spontaneous hydrolysis of the dianion that react s with C-O bond fission and Delta S-not equal=4.2 e.u. The diethylacet al and 2-methyl-G-3-P do not hydrolyze under the same conditions. The reaction occurs with rate-determining intramolecular C-2 deprotonation by the phosphate dianion concerted with its expulsion through the nuc leophilic attack of the carbonylic oxygen. It is proposed that 3-oxete nol is formed as an intermediate that hydrolyzes to glyceraldehyde in a fast step by enforced catalysis of the liberated phosphate dianion. For the hydroxide-catalyzed reaction Delta S-not equal=+32.1 e.u., sug gesting that the virtual transition state is late in the deprotonation step with large contribution of phosphate expulsion. The SIE is 1.85 and orthophosphate appears at the same rate of methylglyoxal. These re sults are consistent with previous conclusion that this reaction proce eds through an E1cb irreversible mechanism with rate-determining C-2 d eprotonation, forming an enediolate intermediate, followed by the expu lsion of phosphate trianion.