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Papers In Press, published online ahead of print October 16, 2003
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Department of Physiology/6085, University of Pennsylvania, Philadelphia, PA 19104-6085
Corresponding Author: cbaron{at}mail.med.upenn.edu
We developed a method for extracting raft-like, liquid-ordered membranes from the particulate fraction prepared from porcine trachealis smooth muscle. This fraction, which contains most of the plasma membrane in this tissue, was homogenized in the presence of cold 0.5% Triton X-100. After centrifugation, membranes containing high contents of sphingomyelin (SM) and cholesterol and low phosphatidylcholine contents remained in the pellet. 35 mM octyl glucoside extracted 75% of these membranes from the Triton X-100-resistant pellet. These membranes had low buoyant densities and accounted for 28% of the particulate fraction lipid. Their lipid composition, 22% SM, 60% cholesterol, 11% PE, 8% PC, <1% PI, and co-isolation with 5'-nucleotidase and caveolin-1 suggest they are liquid-ordered membranes. We compared characteristics of OG and Triton X-100 extractions of the particulate fraction. In contrast to Triton X-100 extractions, membranes released from the particulate fraction by OG were mainly collected in low buoyant fractions at densities ranging from 1.05 to 1.11 g/ml and had phospholipid and cholesterol contents consistent with a mixture of liquid-ordered and liquid-disordered membranes. Thus: OG extraction of apparent liquid-ordered membranes from Triton X-100-resistant pellets was not due to selective extraction of these membranes. Low buoyant density appears not to be unique for liquid-ordered membranes.
Revised on October 13, 2003
Accepted on October 13, 2003
Smooth muscle raft-like membranes
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