PURIFICATION AND CHARACTERIZATION OF OUABAIN-BINDING PROTEIN IN HUMANPLASMA

Ouabainlike factors are thought to be a kind of important modulators o f salt and water metabolism in essential hypertension. We purified the binding-protein of ouabain (OBP) from human plasma. The amino-termina l sequence of OBP from human plasma, (NH2-TLGQPREPQVYTLPPXREEM-), indi cated that OBP is the carboxy-terminal fragment (14.4 kDa by SDS-PAGE) from T218 of IgG2 heavy chain and from A221 of the IgG1 heavy chain c onstant region. Moreover, plasmin-cleaved Fc fragment (pFc) of IgG pos sessed the ouabain-binding activity by the gel-filtration method of pF c and authentic ouabain mixture, whereas neither intact, aggregate, no r papain-cleaved Fc fragment did. The amino-terminal sequence of pFc w as NH2-THTXPPXPAPELLGGPXVFL-, and this sequence corresponded to the T1 05 to L125 fragment of the IgG(1) heavy chain constant region. The gro wth of cultured THP-1 cells were arrested in the dose-dependent manner by ouabain, which was inhibited by the addition of 20 mu g/mL of pFc. These results suggested that plasmin-cleaved Fc of human IgG is one o f the binding protein of ouabain/ouabainlike factor(s) in human plasma .