THE FREE-ENERGY FOR HYDROLYSIS OF A MICROTUBULE-BOUND NUCLEOTIDE TRIPHOSPHATE IS NEAR ZERO - ALL OF THE FREE-ENERGY FOR HYDROLYSIS IS STORED IN THE MICROTUBULE LATTICE
M. Caplow et al., THE FREE-ENERGY FOR HYDROLYSIS OF A MICROTUBULE-BOUND NUCLEOTIDE TRIPHOSPHATE IS NEAR ZERO - ALL OF THE FREE-ENERGY FOR HYDROLYSIS IS STORED IN THE MICROTUBULE LATTICE, The Journal of cell biology, 127(3), 1994, pp. 779-788
The standard free energy for hydrolysis of the GTP analogue guanylyl-(
a,b)-methylene-diphosphonate (GMPCPP), which is -5.18 kcal in solution
, was found to be -3.79 kcal in tubulin dimers, and only -0.90 kcal in
tubulin subunits in microtu- bules. The near-zero change in standard
free energy for GMPCPP hydrolysis in the microtubule indicates that th
e majority of the free energy potentially available from this reaction
is stored in the microtubule lattice; this energy is available to do
work, as in chromosome movement. The equilibrium constants described h
ere were obtained from video microscopy measurements of the kinetics o
f assembly and disassembly of GMPCPP-microtubules and GMPCP-microtubul
es. It was possible to study GMPCPP-microtubules since GMPCPP is not h
ydrolyzed during assembly. Microtubules containing GMPCP were obtained
by assembly of high concentrations of tubulin-GMPCP subunits, as well
as by treating tubulin-GMPCPP-microtubules in sodium (but not potassi
um) Pipes buffer with glycerol, which reduced the halftime for GMPCPP
hydrolysis from >10 h to similar to 10 min. The rate for tubulin-GMPCP
P and tubulin-GMPCP subunit dissociation from microtubule ends were fo
und to be about 0.65 and 128 s(-1), respectively. The much faster rate
for tubulin-GMPCP subunit dissociation provides direct evidence that
microtubule dynamics can be regulated by nucleotide triphosphate hydro
lysis.