Binding of prothrombin and its fragment 1 to phospholipid membranes studied by the solvent relaxation technique.
The phospholipid headgroup mobility of small unilamellar vesicles composed of different mixtures of phosphatidyl-L-serine (PS) and phosphatidylcholine is characterized by the solvent relaxation behavior of the polarity sensitive dyes 6-propionyl-2-(dimethylamino)naphthalene (Prodan) and 6-palmitoyl-2-[trimethylammoniumethyl]-methylamino]naphthalene chloride (Patman). If the PS content exceeds 10%, the addition of calcium leads to a substantial deceleration of the solvent relaxation of both dyes, indicating the formation of Ca(PS)2 complexes. Addition of prothrombin and its fragment 1 leads to a further decrease of the headgroup mobility, as explained by the binding of more than two PS-molecules by a single protein molecule. Prodan monitors the outermost region of the bilayer and it clearly distinguishes between the binding of prothrombin and its fragment 1. The deeper incalated Patman does not distinguish between both proteins. The validity of the solvent relaxation technique for the investigation of the membrane binding of peripheral proteins is demonstrated by the studies of prothrombin induced changes in the steady-state fluorescence anisotropies of 1,6-diphenyl-1,3, 5-hexatriene.
Hutterer
R
R
Institute for Physical Chemistry, University of Würzburg, D-97074 Würzburg, Germany.
Schneider
F W
FW
Hermens
W T
WT
Wagenvoord
R
R
Hof
M
M
D015081:2-Naphthylamine / Q000031:analogs & derivatives; D002118:Calcium / Q000737:chemistry; D005456:Fluorescent Dyes; D008051:Lipid Bilayers / Q000737:chemistry*; D010169:Palmitic Acids; D010446:Peptide Fragments / Q000737:chemistry*; D010713:Phosphatidylcholines / Q000737:chemistry; D010718:Phosphatidylserines / Q000737:chemistry; D010743:Phospholipids / Q000737:chemistry*; D011498:Protein Precursors / Q000737:chemistry*; D011516:Prothrombin / Q000737:chemistry*; D012997:Solvents; D013997:Time Factors
D005456:Fluorescent Dyes; D008051:Lipid Bilayers; D010169:Palmitic Acids; D010446:Peptide Fragments; D010713:Phosphatidylcholines; D010718:Phosphatidylserines; D010743:Phospholipids; D011498:Protein Precursors; D012997:Solvents; C103351:patman; C038517:prodan; C029733:prothrombin fragment 1; D011516:Prothrombin; D015081:2-Naphthylamine; D002118:Calcium