Role of γ carboxyglutamic acid. Cation specificity of prothrombin and factor X phospholipid binding

Gary L Nelsestuen, M. Broderius, G. Martin

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119 Scopus citations


Divalent cations are required for two roles in prothrombin phospholipid interaction. The first role, catalysis of a prothrombin protein transition, has a reaction half life of 100 min at 0°C and is a prerequisite to phospholipid binding. The binding sites required for the transition have a very low cation specificity. All di and trivalent cations tested were effective in this role with the exception of beryllium. Barium catalyzed the transition but only at high concentrations (6.6 mM was required for half reaction). Blood clotting Factor X, another γ carboxyglutamic acid containing protein, also undergoes a cation catalyzed protein transition which is a prerequisite to Factor X phospholipid binding. In both proteins, the transition can be monitored by a decrease in the protein's intrinsic fluorescence. Compared to prothrombin, the Factor X transition occurs much more rapidly, has a somewhat greater specificity for cations, and requires higher concentrations of cations. This indicates that the cation binding sites provided by γ carboxyglutamic acid are not completely uniform in all proteins. The second role of divalent cations in prothrombin phospholipid interaction is in the actual protein phospholipid binding. This interaction was studied by protein fluorescence quenching resulting from excitation energy transfer to a chromophore attached to the phospholipid membrane. Only strontium and barium satisfactorily replaced calcium in this role. A number of other cations form protein phospholipid complexes but of the wrong structure. These cations inhibit the prothrombinase complex (Factor X(a), calcium, phospholipid, Factor V). The cation specificity for Factor X phospholipid binding is the same as for prothrombin except that higher concentrations of cations are required. Factor X(a) (generated by action of Russell's viper venom on Factor X) displayed the same calcium requirements for the protein transition and phospholipid interaction as Factor X. The cation requirements of the prothrombinase complex correlate with the cation requirements of prothrombin and Factor X phospholipid binding. Strontium is the only cation that will singly replace calcium. Barium is ineffective alone because the concentrations required to catalyze the protein transitions cause precipitation of the phospholipid. Combination of certain other cations with barium will, however, substitute for calcium. The other cations (specifically magnesium or manganous ion) catalyze the protein transitions and barium forms the correct protein phospholipid complexes.

Original languageEnglish (US)
Pages (from-to)6886-6893
Number of pages8
JournalJournal of Biological Chemistry
Issue number22
StatePublished - 1976


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