Synthesis of phospholipid-based detergents and their effects on the Ca(2+)-ATPase of sarcoplasmic reticulum.
Phospholipid molecules containing a cholate or hemisuccinylcholate moiety at the 2-position were synthesized as possible detergents for solubilization of membrane proteins. 1-Myristoleoyl-2-cholyl-sn-glycero-3- phosphatidylcholine ((C14:1,cholyl)PC) was found to solubilize sarcoplasmic reticulum vesicles at concentrations above its cmc of ca. 4 micrograms/ml. Effects of (C14:1,cholyl)PC on the activity of the Ca(2+)-ATPase of sarcoplasmic reticulum were complex, as for other detergents. High concentrations (0.2 mg/ml) of (C14:1,cholyl)PC were able to displace phospholipids from the lipid/protein interface of the ATPase. Although under these conditions the activity of the ATPase was low, the ATPase was not denatured since activity could be regained by displacement of (C14:1,cholyl)PC with the detergent C12E8. 1-oleoyl-2-cholyl-sn-glycero-3-phosphatidylcholine ((C18:1,cholyl)PC) was found to have a very low water solubility, but this could be increased by the introduction of a hemisuccinyl group to give 1-oleoyl-2-(3 alpha-hemisuccinyl)cholyl-sn-glycero-3-phosphatidylcholine ((C18:1,cholylCOOH)PC). This was able to solubilize and delipidate the Ca(2+)-ATPase; the ATPase was stable in (C18:1,cholylCOOH)PC for long periods of time.
Ding
J
J
Department of Biochemistry, University of Southampton, UK.
East
J M
JM
Lee
A G
AG
D000818:Animals; D000252:Calcium-Transporting ATPases / Q000302:isolation & purification / Q000378:metabolism*; D003902:Detergents / Q000138:chemical synthesis* / Q000494:pharmacology; D004795:Enzyme Stability; D007202:Indicators and Reagents; D007700:Kinetics; D018482:Muscle, Skeletal / Q000201:enzymology*; D010713:Phosphatidylcholines / Q000138:chemical synthesis* / Q000494:pharmacology; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D013050:Spectrometry, Fluorescence
D003902:Detergents; D007202:Indicators and Reagents; D010713:Phosphatidylcholines; D000252:Calcium-Transporting ATPases