title: On the kinetics of adsorption and two-dimensional self-assembly of annexin A5 on supported lipid bilayers.
abstract: Annexin A5 is a protein that binds to membranes containing negatively charged phospholipids in a calcium-dependent manner. We previously found that annexin A5 self-assembles into two-dimensional (2D) crystals on supported lipid bilayers (SLBs) formed on mica while a monolayer of disordered trimers is formed on SLBs on silica. Here, we investigated in detail and correlated the adsorption kinetics of annexin A5 on SLBs, supported on silica and on mica, with the protein's 2D self-assembly behavior. For this study, quartz crystal microbalance with dissipation monitoring and ellipsometry were combined with atomic force microscopy. We find, in agreement with previous studies, that the adsorption behavior is strongly dependent on the concentration of dioleoylphosphatidylserine (DOPS) in the SLB and the calcium concentration in solution. The adsorption kinetics of annexin A5 are similar on silica-SLBs and on mica-SLBs, when taking into account the difference in accessible DOPS between silica-SLBs and mica-SLBs. In contrast, 2D crystals of annexin A5 form readily on mica-SLBs, even at low protein coverage (< or =10%), whereas they are not found on silica-SLBs, except in a narrow range close to maximal coverage. These results enable us to construct the phase diagram for the membrane binding and the states of 2D organization of annexin A5. The protein binds to the membrane in two different fractions, one reversible and the other irreversible, at a given calcium concentration. The adsorption is determined by the interaction of protein monomers with the membrane. We propose that the local membrane environment, as defined by the presence of DOPS, DOPC, and calcium ions, controls the adsorption and reversibility of protein binding.
authors.affiliation: Laboratoire d'Imagerie Moléculaire et NanoBioTechnologie, IECB, UMR-CNRS 5471, Université Bordeaux 1, 33607 Pessac Cedex, France.
authors.forename: Ralf P
authors.initials: RP
authors.lastname: Richter
authors.forename: Joséphine Lai Kee
authors.initials: JL
authors.lastname: Him
authors.forename: Béatrice
authors.initials: B
authors.lastname: Tessier
authors.forename: Céline
authors.initials: C
authors.lastname: Tessier
authors.forename: Alain R
authors.initials: AR
authors.lastname: Brisson
mesh_terms: D000327:Adsorption; D000818:Animals; D017304:Annexin A5 / Q000737:chemistry*; D002118:Calcium / Q000737:chemistry; D003460:Crystallization; D018360:Crystallography, X-Ray; D019281:Dimerization; D007700:Kinetics; D008051:Lipid Bilayers / Q000737:chemistry*; D018625:Microscopy, Atomic Force; D010713:Phosphatidylcholines / Q000737:chemistry; D010718:Phosphatidylserines / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry; D012825:Silicon / Q000737:chemistry; D013997:Time Factors
chemical_list: D017304:Annexin A5; D008051:Lipid Bilayers; D010713:Phosphatidylcholines; D010718:Phosphatidylserines; D011994:Recombinant Proteins; C042511:1,2-dioleoylphosphatidylserine; C017251:1,2-oleoylphosphatidylcholine; D002118:Calcium; D012825:Silicon