FACTOR-VA MEMBRANE INTERACTION IS MEDIATED BY 2 REGIONS LOCATED ON THE LIGHT-CHAIN OF THE COFACTOR

Factor Va was incubated with 1-azidopyrene, a fluorescent lipophilic p robe, in the presence of phospholipid vesicles composed of various pro portions of phosphatidylcholine (PC) and phosphatidylserine (PS). The majority of the label was associated with the light chain of factor Va . The light chain was found to be labeled in the presence of phospholi pid vesicles containing either 100% PC or 100%PS. After cleavage by fa ctor Xa and incubation with PC/PS vesicles composed of 75% PC and 25% PS, label was found both on the M(r) = 30 000 fragment, derived-from t he NH2-terminal portion of the bovine factor Va light chain (residues 1537-1752), and on the M(r) = 46 000/48 000 carboxyl-terminal fragment of the factor Va light chain (residues 1753-2183). The M(r) = 46 000/ 48 000 fragment incorporated I-azidopyrene independent of the phosphol ipid composition, while label incorporation into the M(r) = 30 000 fra gment required phospholipid vesicles containing PC. No labeling of the M(r) = 30 000 fragment was observed with phospholipid vesicles compos ed of 100% PS. The label incorporation into the two portions of the mo lecule was found to be independent of the ionic strength in the presen ce of phospholipid vesicles containing 75% PC and 25% PS. In contrast, the labeling of the M(r) = 46 000/48 000 fragment with phospholipid v esicles composed of 1 00% PS was ionic strength dependent. These data suggest that two regions of factor Va light chain interact with the li pid bilayer and have different requirements for interaction: the bindi ng site located on the M(r) = 30 000 fragment of the cofactor (A3 doma in) interacts with phospholipid vesicles containing neutral phospholip id and is most likely hydrophobic in nature whereas the binding site l ocated on the M(r) = 46 000/48 000 carboxyl-terminal fragment (C1-C2 d omains ) interacts with membranes composed of anionic and neutral phos pholipid and displays partly ionic binding characteristics.