Thermodynamic properties of the kinesin neck-region docking to the catalytic core.
Kinesin motors move on microtubules by a mechanism that involves a large, ATP-triggered conformational change in which a mechanical element called the neck linker docks onto the catalytic core, making contacts with the core throughout its length. Here, we investigate the thermodynamic properties of this conformational change using electron paramagnetic resonance (EPR) spectroscopy. We placed spin probes at several locations on the human kinesin neck linker and recorded EPR spectra in the presence of microtubules and either 5'-adenylylimidodiphosphate (AMPPNP) or ADP at temperatures of 4-30 degrees C. The free-energy change (DeltaG) associated with AMPPNP-induced docking of the neck linker onto the catalytic core is favorable but small, about 3 kJ/mol. In contrast, the favorable enthalpy change (DeltaH) and unfavorable entropy change (TDeltaS) are quite large, about 50 kJ/mol. A mutation in the neck linker, V331A/N332A, results in an unfavorable DeltaG for AMPPNP-induced zipping of the neck linker onto the core and causes motility defects. These results suggest that the kinesin neck linker folds onto the core from a more unstructured state, thereby paying a large entropic cost and gaining a large amount of enthalpy.
Rice
S
S
Department of Cellular and Molecular Pharmacology, University of California, San Francisco 94143, USA.
Cui
Y
Y
Sindelar
C
C
Naber
N
N
Matuska
M
M
Vale
R
R
Cooke
R
R
D001665:Binding Sites; D020134:Catalytic Domain; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods*; D004735:Energy Transfer; D006801:Humans; D016547:Kinesins / Q000737:chemistry* / Q000235:genetics; D008870:Microtubules / Q000737:chemistry*; D020409:Molecular Motor Proteins / Q000737:chemistry*; D009038:Motion; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D013113:Spin Labels / Q000138:chemical synthesis; D013314:Stress, Mechanical; D013696:Temperature
D020409:Molecular Motor Proteins; D011994:Recombinant Proteins; D013113:Spin Labels; D016547:Kinesins