Annexin A4 binding to anionic phospholipid vesicles modulated by pH and calcium.
Annexin A4 belongs to a class of Ca(2+)-binding proteins for which different functions in the cell have proposed, e.g. involvement in exocytosis and in the coagulation process. All these functions are related to the ability of the annexins to bind to acidic phospholipids. In this study the interaction of annexin A4 with large unilamellar vesicles (LUV) prepared from phosphatidylserine (PS) or from phosphatidic acid (PA) is investigated at neutral and acidic pH. Annexin A4 strongly binds to either lipid at acidic pH, whereas at neutral pH only weak binding to PA and no binding to PS occurs. Addition of 40 microM Ca(2+) leads to a strong binding to the lipids also at neutral pH. This is caused by the different electric charge of the protein below and above its isoelectric point. Binding of annexin A4 induces dehydration of the vesicle surface. The strength of the effects is much greater at pH 4 than at pH 7.4. At pH 7.4 annexin A4 reduces the Ca(2+)-threshold concentration necessary to induce fusion of PA LUV. The Ca(2+) induced fusion of PS LUV is not affected by annexin A4 at pH 7.4. At pH 4 annexin A4 induces fusion of either vesicles without Ca(2+). Despite the low binding extents at neutral pH annexin A4 induces a Ca(2+) independent leakage of PS- or PA-LUV. The leakage extent is increased at acidic pH. From the data two suggestions are made: (1) At pH 4 annexin A4 (at least partially) penetrates into the bilayer in contrast to the preferred location at the vesicle surface at neutral pH. The conformation of annexin A4 seems to be different at the two conditions. (2) At neutral pH, Annexin A4 seems to be able to bind two PA vesicles simultaneously; however, only one PS vesicle at the same time. This behavior might be related to a recently described double Ca(2+) binding site, which appears to be uniquely suited for PS.
Zschörnig
Olaf
O
Institute of Medical Physics and Biophysics, University of Leipzig, Leipzig, Germany. olaf.zschoernig@medizin.uni-leipzig.de
Opitz
Frank
F
Müller
Matthias
M
D000838:Anions; D017314:Annexin A4 / Q000737:chemistry*; D001665:Binding Sites; D002118:Calcium / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D008081:Liposomes / Q000737:chemistry*; D008956:Models, Chemical; D010743:Phospholipids / Q000737:chemistry*; D011485:Protein Binding
D000838:Anions; D017314:Annexin A4; D008081:Liposomes; D010743:Phospholipids; D002118:Calcium