Amphipathic Helices Support Function of Blood Coagulation Factor IXa

Blood coagulation factor IXa gains proteolytic efficiency upon binding to a phospholipid membrane. We have found that an amphipathic, membrane-binding peptide from the C2 domain of factor VIII, fVIII2303 - 23, enhances proteolytic efficiency of factor IXa in the absence of phospholipid membranes. This enhancement is the result of a reduction in the K M for the substrate, factor X, with little effect on the k cat. Enhanced function requires interaction of the γ-carboxyglutamic acid (Gla) domains of factor IXa and factor X since (i) a synthetic peptide comprising the Gla domain of factor IXa and antibodies directed to the Gla domain of factor IXa inhibit this acceleration, (ii) the acceleration is Ca(II) dependent, and (iii) conversion of Gla-domainless factor X is not affected by the presence of fVIII2303 - 23. The effect of fVIII2303 - 23 on factor IXa parallels the enhanced function produced by phosphatidylserine-containing bilayers, and fVIII2303 - 23 does not further enhance function of factor IXa when phospholipid vesicles are present. The critical feature of fVIII2303 - 23 is apparently its amphipathic helix-forming structure [Gilbert, G. E., and Baleja, J. D. (1995) Biochemistry 34, 3022−3031] because other α-helical peptides such as a homologous peptide from the C2 domain of factor V and melittin have similar effects. Diastereomeric analogues of fVIII2303 - 23 and melittin, which have reduced helical content, do not support factor IXa activity. A truncated peptide of fVIII2303 - 23 with three C-terminal residues deleted retains α-helical content but loses capacity to enhance factor X cleavage, suggesting that a minimum length of α-helix is required. Although these results probably do not illuminate the physiologic function of the factor VIII peptide corresponding to fVIII2303 - 23, they demonstrate a novel, membrane-mimetic role of amphipathic helical peptides in supporting function of factor IXa.