The relationship between the binding of ATP and calcium to annexin IV. Effect of nucleotide on the calcium-dependent interaction of annexin with phosphatidylserine

Abstract

With the use of ATP analogues, we have found that porcine liver annexin (Anx) IV can be covalently labelled with 8-azido[γ-³²P]-ATP In the presence of Ca²⁺ (K_d 4.2 μM) and that the labelling is prevented by asolectin/cholesterol liposomes or chelation of calcium ions. On the other hand, non-covalent binding of 2 -(or 3)-0-(2,4,6-trinitrophenyl)adenoslne 5-triphosphate (TNP-ATP) to AnxlV occurs optimally in the presence of liposomes and Ca²⁺ (K_d 7 μM). These observations were further confirmed by the results of intrinsic fluorescence quenching of AnxlV with various nucleotides, suggesting the existence of a relationship between Ca²⁺, phospholipid- and ATP-binding sites within the annexin molecule. The Interaction of AnxIV with nucleotides does not significantly affect its In vitro properties concerning the binding to phosphatidylserine (PS) monolayers.

Key Words:

• annexin IV
• 8-azido-ATP
• TNP-ATP
• fluorescence quenching
• PS monolayer