Abstract
Synaptotagmin I has two tandem Ca2+-binding C2 domains, which are essential for fast synchronous synaptic transmission in the central nervous system. We have solved four crystal structures of the C 2B domain, one of them in the cation-free form at 1.50 Å resolution, two in the Ca2+-bound form at 1.04 Å (two bound Ca2+ ions) and 1.65 Å (three bound Ca2+ ions) resolution and one in the Sr2+-bound form at 1.18 Å (one bound Sr2+ ion) resolution. The side chains of four highly conserved aspartic acids (D303, D309, D363, and D365) and two main chain oxygens (M302:O and Y364:O), together with water molecules, are in direct contact with two bound Ca2+ ions (sites 1 and 2). At higher Ca2+ concentrations, the side chain of N333 rotates and cooperates with D309 to generate a third Ca2+ coordination site (site 3). Divalent cation binding sites 1 and 2 in the C2B domain were previously identified from NMR NOE patterns and titration studies, supplemented by site-directed mutation analysis. One difference between the crystal and NMR studies involves D371, which is not involved in coordination with any of the identified Ca2+ sites in the crystal structures, while it is coordinated to Ca2+ in site 2 in the NMR structure. In the presence of Sr2+, which is also capable of triggering exocytosis, but with lower efficiency, only one cation binding site (site 1) was occupied in the crystallographic structure.
Original language | English (US) |
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Pages (from-to) | 2665-2672 |
Number of pages | 8 |
Journal | Protein Science |
Volume | 13 |
Issue number | 10 |
DOIs | |
State | Published - Oct 2004 |
Externally published | Yes |
Keywords
- CB domain
- Calcium binding
- Strontium binding
- Synaptotagmin I
- X-ray crystallography