GTP hydrolysis by tubulin occurs with water oxygen incorporation into inorganic phosphate

Abstract

Tubulin assembly was conducted in [ 18O]H 2O and the resulting mixture of GTP, GDP and P i was examined by 31P-NMR. Two P i signals, separated by about 0.02 ppm, were observed. By combining this mixture with a solution of P i containing all five possible 16O and 18O isotopomers of P i, it was shown that the two signals were due to [ 16O 4]- and [ 16O 3, 18O]P i. The amount of 18O incorporated into the P i was that expected if the hydrolysis of GTP during tubulin assembly occurs with cleavage of the γ-phosphorus-bridge oxygen bond.