NMR measurement of the off rate from the first calcium-binding site of the synaptotagmin I C2A domain.
The off rate from the first calcium-binding site of the C2A domain of synaptotagmin I, a putative calcium receptor in neurotransmitter release, has been determined by 15N-nuclear magnetic resonance relaxation dispersion measurements. The exchange rate was obtained by fitting the dependence of the transverse relaxation rates on the interval between 180 degrees pulses in relaxation-compensated CPMG experiments at 3.2 microM calcium concentration. The measured k(ex) is 2.0x10(3) x s(-1). The calcium on rate of 3.5+/-1x10(7) x s(-1), determined from the measured off rate and the dissociation constant (5.3x10(-5) M), is close to the diffusion limit. These results are consistent with the proposed role of synaptotagmin I as a calcium sensor in release, but suggest that additional factors may help to accelerate the diffusion of Ca2+ to the sensor.
Millet
Oscar
O
Departament de Química Orgànica, Universitat de Barcelona, Martí i Franquès 1-11, 08028, Barcelona, Spain.
Bernadó
Pau
P
Garcia
Jesús
J
Rizo
Josep
J
Pons
Miquel
M
D001665:Binding Sites; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling; D002135:Calcium-Binding Proteins; D066298:In Vitro Techniques; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000378:metabolism*; D008959:Models, Neurological; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000378:metabolism*; D018377:Neurotransmitter Agents / Q000378:metabolism; D017434:Protein Structure, Tertiary; D050863:Synaptotagmin I; D050857:Synaptotagmins
D002135:Calcium-Binding Proteins; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; D018377:Neurotransmitter Agents; D050863:Synaptotagmin I; D050857:Synaptotagmins; D002118:Calcium