title: Membrane modulates affinity for calcium ion to create an apparent cooperative binding response by annexin a5.
abstract: Isothermal titration calorimetry was used to characterize the binding of calcium ion (Ca²⁺) and phospholipid to the peripheral membrane-binding protein annexin a5. The phospholipid was a binary mixture of a neutral and an acidic phospholipid, specifically phosphatidylcholine and phosphatidylserine in the form of large unilamellar vesicles. To stringently define the mode of binding, a global fit of data collected in the presence and absence of membrane concentrations exceeding protein saturation was performed. A partition function defined the contribution of all heat-evolving or heat-absorbing binding states. We find that annexin a5 binds Ca²⁺ in solution according to a simple independent-site model (solution-state affinity). In the presence of phosphatidylserine-containing liposomes, binding of Ca²⁺ differentiates into two classes of sites, both of which have higher affinity compared with the solution-state affinity. As in the solution-state scenario, the sites within each class were described with an independent-site model. Transitioning from a solution state with lower Ca²⁺ affinity to a membrane-associated, higher Ca²⁺ affinity state, results in cooperative binding. We discuss how weak membrane association of annexin a5 prior to Ca²⁺ influx is the basis for the cooperative response of annexin a5 toward Ca²⁺, and the role of membrane organization in this response.
authors.affiliation: Department of Chemistry and Biochemistry, University of Minnesota Duluth, Duluth, MN, USA.
authors.forename: Jacob W
authors.initials: JW
authors.lastname: Gauer
authors.forename: Kristofer J
authors.initials: KJ
authors.lastname: Knutson
authors.forename: Samantha R
authors.initials: SR
authors.lastname: Jaworski
authors.forename: Anne M
authors.initials: AM
authors.lastname: Rice
authors.forename: Anika M
authors.initials: AM
authors.lastname: Rannikko
authors.forename: Barry R
authors.initials: BR
authors.lastname: Lentz
authors.forename: Anne
authors.initials: A
authors.lastname: Hinderliter
mesh_terms: D017304:Annexin A5 / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism*; D008563:Membrane Lipids / Q000737:chemistry / Q000378:metabolism; D008954:Models, Biological; D011485:Protein Binding
chemical_list: D017304:Annexin A5; D008563:Membrane Lipids; D002118:Calcium