PDBsum entry 2cm6

Protein transport

PDB id: 2cm6

Contents

Protein chains

149 a.a. *

Ligands

PO4

Metals

_CA ×4

Waters ×76

* Residue conservation analysis

PDB id:

2cm6

Name:

Protein transport

Title:

Crystal structure of the c2b domain of rabphilin3a

Structure:

Rabphilin-3a. Chain: a, b. Fragment: c2b domain and linker, residues 519-684. Synonym: exophilin-1. Engineered: yes

Source:

Rattus norvegicus. Rat. Organism_taxid: 10116. Expressed in: escherichia coli. Expression_system_taxid: 511693.

Biol. unit:

Monomer (from PDB file)

Resolution:

1.85Å R-factor: 0.198 R-free: 0.268

Authors:

C.Schlicker,P.Montaville,G.M.Sheldrick,S.Becker

Key ref:

P.Montaville et al. (2007). The C2A-C2B linker defines the high affinity Ca(2+) binding mode of rabphilin-3A. J Biol Chem, 282, 5015-5025. PubMed id: 17166855 DOI: 10.1074/jbc.M606746200

Date:

04-May-06 Release date: 04-Dec-06

Protein chains

P47709  (RP3A_RAT) -  Rabphilin-3A from Rattus norvegicus

Seq: 684 a.a.

Struc: 149 a.a.

DOI no: 10.1074/jbc.M606746200

J Biol Chem 282:5015-5025 (2007)

PubMed id: 17166855

The C2A-C2B linker defines the high affinity Ca(2+) binding mode of rabphilin-3A.

P.Montaville, C.Schlicker, A.Leonov, M.Zweckstetter, G.M.Sheldrick, S.Becker.

ABSTRACT

The Ca(2+) binding properties of C2 domains are essential for the function of their host proteins. We present here the first crystal structures showing an unexpected Ca(2+) binding mode of the C2B domain of rabphilin-3A in atomic detail. Acidic residues from the linker region between the C2A and C2B domains of rabphilin-3A interact with the Ca(2+)-binding region of the C2B domain. Because of these interactions, the coordination sphere of the two bound Ca(2+) ions is almost complete. Mutation of these acidic residues to alanine resulted in a 10-fold decrease in the intrinsic Ca(2+) binding affinity of the C2B domain. Using NMR spectroscopy, we show that this interaction occurred only in the Ca(2+)-bound state of the C2B domain. In addition, this Ca(2+) binding mode was maintained in the C2 domain tandem fragment. In NMR-based liposome binding assays, the linker was not released upon phospholipid binding. Therefore, this unprecedented Ca(2+) binding mode not only shows how a C2 domain increases its intrinsic Ca(2+) affinity, but also provides the structural base for an atypical protein-Ca(2+)-phospholipid binding mode of rabphilin-3A.

Selected figure(s)

Figure 1.
FIGURE 1. Structure of rabphilin-3A C2B domain fragment 519-684. A, ribbon model of the low Ca^2+ (P2[1]2[1]2) structure. The Ca^2+ ions are presented as orange spheres. All structural figures were generated with PyMOL (pymol.sourceforge.net). B, ribbon model of monomer A (green) of the high Ca^2+ structure (P2[1]). C and D, 2mF[o] - DF[c] electron density maps of the linker region in the low and high (monomer A) Ca^2+ structures, respectively.

Figure 3.
FIGURE 3. Interaction network between the linker region and the CBLs. A, low Ca^2+ structure; B, high Ca^2+ structure; C, superposition with the Ca^2+ ions of the synaptotagmin 1 C2A domain (Protein Data Bank code 1BYN; in yellow). For clarity, the N-terminal linker region is not shown. Hydrogen bonds involving Lys^636 are depicted in green. Ca 1 and Ca 2, Ca^2+ in binding sites 1 and 2, respectively.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2007, 282, 5015-5025) copyright 2007.

Figures were selected by an automated process.