Insight into the location and dynamics of the annexin A2 N-terminal domain during Ca(2+)-induced membrane bridging.
Annexin A2 (AnxA2) is a Ca(2+)- and phospholipid-binding protein involved in many cellular regulatory processes. Like other annexins, it is constituted by two domains: a conserved core, containing the Ca(2+) binding sites, and a variable N-terminal segment, containing sites for interactions with other protein partners like S100A10 (p11). A wealth of data exists on the structure and dynamics of the core, but little is known about the N-terminal domain especially in the Ca(2+)-induced membrane-bridging process. To investigate this protein region in the monomeric AnxA2 and in the heterotetramer (AnxA2-p11)(2), the reactive Cys8 residue was specifically labelled with the fluorescent probe acrylodan and the interactions with membranes were studied by steady-state and time-resolved fluorescence. In membrane junctions formed by the (AnxA2-p11)(2) heterotetramer, the flexibility of the N-terminal domain increased as compared to the protein in solution. In "homotypic" membrane junctions formed by monomeric AnxA2, acrylodan moved to a more hydrophobic environment than in the protein in solution and the flexibility of the N-terminal domain also increased. In these junctions, this domain is probably not in close contact with the membrane surface, as suggested by the weak quenching of acrylodan observed with doxyl-PCs, but pairs of N-termini likely interact, as revealed by the excimer-forming probe pyrene-maleimide bound to Cys8. We present a model of monomeric AnxA2 N-terminal domain organization in "homotypic" bridged membranes in the presence of Ca(2+).
Ayala-Sanmartin
Jesus
J
INSERM U538, CHU Saint-Antoine, Paris F-75012, France. jayala@chusa.jussieu.fr
Zibouche
Mallik
M
Illien
Françoise
F
Vincent
Michel
M
Gallay
Jacques
J
D017306:Annexin A2 / Q000737:chemistry*; D002118:Calcium / Q000737:chemistry*; D008567:Membranes, Artificial; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence
D017306:Annexin A2; D008567:Membranes, Artificial; D002118:Calcium