Structural basis for the evolutionary inactivation of Ca2+ binding to synaptotagmin 4.
The neuronal protein synaptotagmin 1 functions as a Ca(2+) sensor in exocytosis via two Ca(2+)-binding C(2) domains. The very similar synaptotagmin 4, which includes all the predicted Ca(2+)-binding residues in the C(2)B domain but not in the C(2)A domain, is also thought to function as a neuronal Ca(2+) sensor. Here we show that, unexpectedly, both C(2) domains of fly synaptotagmin 4 exhibit Ca(2+)-dependent phospholipid binding, whereas neither C(2) domain of rat synaptotagmin 4 binds Ca(2+) or phospholipids efficiently. Crystallography reveals that changes in the orientations of critical Ca(2+) ligands, and perhaps their flexibility, render the rat synaptotagmin 4 C(2)B domain unable to form full Ca(2+)-binding sites. These results indicate that synaptotagmin 4 is a Ca(2+) sensor in the fly but not in the rat, that the Ca(2+)-binding properties of C(2) domains cannot be reliably predicted from sequence analyses, and that proteins clearly identified as orthologs may nevertheless have markedly different functional properties.
Department of Biochemistry, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, Texas 75390, USA.
Han
H
Dai
Ok-Ho
OH
Shin
Mischa
M
Machius
Diana R
DR
Tomchick
Thomas C
TC
Südhof
Josep
J
Rizo
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000737:chemistry* / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D018360:Crystallography, X-Ray; D019143:Evolution, Molecular; D005089:Exocytosis; D005982:Glutathione Transferase / Q000378:metabolism; D008024:Ligands; D008055:Lipids / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000378:metabolism; D008954:Models, Biological; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000378:metabolism; D009474:Neurons / Q000378:metabolism; D010743:Phospholipids / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D050863:Synaptotagmin I; D050857:Synaptotagmins
D002135:Calcium-Binding Proteins; D008024:Ligands; D008055:Lipids; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; D010743:Phospholipids; D011993:Recombinant Fusion Proteins; D050863:Synaptotagmin I; C494312:Syt1 protein, rat; C494294:Syt4 protein, rat; D050857:Synaptotagmins; D005982:Glutathione Transferase; D002118:Calcium