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J P Segrest

Publications and source records attributed to J P Segrest.

At least 127 records · Page 7Linked to original sources
PubMed1979

Isolation of glycophorin with deoxycholate.

In a previous communication we reported that human erythrocyte glycophorin prepared by the lithium diiodosalicylate phenol procedure contains approximately 10 mol of lithium diiodosalicylate per mol of glycophorin, and further we showed that this bound lithium diiodosalicylate is difficult to remove by detergents or organic solvents (Romans, A.Y. and Segrest, J.P. (1978) Biochim. Biophys. Acta 511, 297-301). In the present communication we report an alternative purification procedure for glycophorin in which sodium deoxycholate is substituted for lithium diiodosalicylate; the sodium deoxycholate is subsequently removed by gel filtration. Utilizing this procedure, 25-30 mg glycophorin are obtained per gram of lyophilized erythrocyte ghosts. The glycophorin prepared by the sodium deoxycholate procedure, after a single gel filtration step, contains less than 1 mol of sodium deoxycholate per mol glycophorin and is colorless compared with glycophorin prepared by the lithium diiodosalicylate procedure, which has a distint reddish-brown cast.

Chromatography, Gel↗
PubMed1978

Binding of lithium diiodosalicylate to glycophorin.

Glycophorin was purified from human erythrocyte ghosts by the lithium diiodosalicylate -phenol procedure utilizing 125I-labeled lithium diiodosalicylate. The glycophorin preparation was found to contain 8.9 +/- 2.1 mol lithium diiodosalicylate per mol glycophorin. This bound lithium diiodosalicylate cannot be removed by extensive washings with a variety of polar organic solvents nor by treatment with the detergent, sodium deoxycholate. Further, the hydrophobic peptide produced from glycophorin by trypsin digestion contained 3.4 mol lithium diiodosalicylate per mol peptide.

Erythrocyte Membrane↗
PubMed1977

Amphipathic helixes and plasma lipoproteins: thermodynamic and geometric considerations.

In this paper analyses are made of the thermodynamic and geometric properties of the predicted association between amphipathic helixes and phospholipid vesicles. From thermodynamic considerations it is proposed that a major driving force for such an association is the negative free energy gained by the transfer of a number of hydrophobic residues (contained within the non-polar faces of amphipathic helixes), from water to the interior of a phospholipid bilayer. The mechanism proposed is that in the aqueous state a potentially amphipathic sequence forms a non-helical hydrophobic patch on the surface of the apolipoprotein. Formation of an amphipathic helix and simultaneous burial of the hydrophobic residues in the surface of a phospholipid bilayer provides the driving force for lipid association. From this model an estimate of the upperlimit for the hydrophobically driven free energy of lipid association (-40-65 kcal/mol) is calculated for the 4 apolipoproteins with known sequences. On the basis of geometrical considerations a model for an intermediate state of high density lipoprotein (HDL) synthesis is proposed. This model consists of a cholesterol-containing phospholipid bilayer disc whose 'naked" hydrophobic edges are shielded from the aqueous phase by amphipathic helixes of the apolipoproteins. Exposure of these 'bicycle tire" miscelles to the enzyme lecithin : cholesterol acyl transferase (LCAT) is postulated to result in the formation of mature spherical HDL particles with cholesteryl ester forming a neutral lipid core.

Apolipoproteins↗
PubMed1977

Interaction of the lipid-associating domain of glycophorin with phospholipid bilayers.

The MN-blood group bearing glycoprotein of the human red cell membrane, glycophorin, has been shown to traverse the membrane in vivo such that the NH2-terminal portion is exposed to the external, and the COOH-terminal portion exposed to the cytoplasmic, mileiu. The intervening 23 residue lipid-associating domain (LAD) of glycophorin has been shown to have a unique overall hydrophobicity. The LAD of glycophorin can be obtained intact within an aqueous insoluble tryptic peptic, T(is). Under appropriate conditions (the reaction not being spontaneous), T(is) can be associated with phospholipid bilayers. Freeze fracture studies of T(is): phospholipid vesicles suggest that T(is) forms multimeric torus-shaped intramembranous structures 80 A in diameter with n greater than 4. The plot of T(is) concentration versus multimer density suggests there is a critical multimer concentration (CMC) for T(is) in phospholipid bilayers (L/P = 200/1). Various physico-chemical techniques such as pyrene fluorescence spectroscopy, differential scanning calorimetry, and ionic permeability were used to investigate the T(is)-lipid association. Results indicate that this system is an excellent one in which to study the boundary lipid phenomenon. In addition, T(is) association with lipid bilayers is being correlated with the natural state of glycophorin in the red cell membrane.

Chemical Phenomena↗
PubMed1976

Amyloid A: amphipathic helixes and lipid binding.

Polypeptide segments, composed of alpha helixes with specific surface topography termed amphipathic helixes, have been proposed as the basic lipid-associating domains of apolipoproteins from the plasma lipoproteins. A computer search for proteins having sequences that could form amphipathic helixes indicated that amyloid A, a pathologically occurring protein usually associated with "secondary" amyloidosis, also contained amphipathic helixes. In studies reported here, amyloid A is shown to associate spontaneously with phospholipid vesicles with the following results: (a) the formation of a protein-lipid complex isolated by equilibrium density gradient ultracentrifugation, (b) a 100% increase in alpha helicity as measured by circular dichroism, (c) a 9-nm shift in the fluorescence maximum due to the single tryptophan residue located in the amphipathic region, indicating the tryptophan is moving from a polar to a nonpolar environment, and (d) the formation of stacked disk-like protein-lipid complexes as visualized by negative stain electron microscopy. The temperature dependence of the circular dichroic spectrum of the amyloid A-phospholipid complex suggests that the complex is formed by insertion of protein between the fatty acyl chains of the lipid. These findings suggest that the amphipathic helix is an important structural unit in lipid-associating proteins and that this unit can be recognized on the basis of its amino acid sequence. In addition, these studies have implications for the origin and function of amyloid A protein.

Amyloid↗
PubMed1975

The effects of the membrane-penetrating polypeptide segment of the human erythrocyte MN-glycoprotein on the permeability of model lipid membranes.

The insoluble peptide, T(is), prepared by trypsin hydrolysis of the MN-glycoprotein (glycophorin) of the human erythrocyte has been incorporated into phospholipid membranes in the form of liposomes and black lipid membranes. The permeability of liposome membranes to 42K+ and of black lipid membranes to water and ions is increased significantly by the presence of the T(is) peptide. Electrophoresis measurements indicate that these effects are not due to the T(is) peptide carrying a net charge. The results suggest that the peptide causes local disordering of the bilayer membrane structures. This is considered in the light of findings published elsewhere: that the MN-glycoprotein penetrates through the cell membrane via a non-polar segment of its polypeptide chain, which is contained intact within the T(is) peptide; that the T(is) peptide is partially helical when associated with phospholipid and forms multimeric 8.0 nm structures within the hydrophobic plane of phospholipid bilayers.

Biological Transport↗
PubMed1975

The mechanism of lipid-binding by plasma lipoproteins.

In this article, we review the current state of knowledge about the structure of human plasma high density lipoproteins. Recent advances in this field, including amino acid sequencing and lipid-binding studies, suggest that the plasma lipoproteins have specific sites with special structural features that account for their ability to bind and transport lipid. A hypothesis which describes the structure of these lipid-binding sites and their interaction with lipid is presented.

Amino Acids↗
PubMed1974

Association of the membrane-penetrating polypeptide segment of the human erythrocyte MN-glycoprotein with phospholipid bilayers. I. Formation of freeze-etch intramembranous particles.

The membrane-penetrating segment of the surface MN-glycoprotein of the human erythrocyte is contained intact within the tryptic peptide T(is). We report here on the association of this peptide with hydrated phospholipid vesicles. Under these conditions 80 A intramembranous particles, as seen by freeze-etch electron microscopy, are produced that are similar in size to those seen in the native erythrocyte membrane. These particles increase in number as a linear function of T(is) concentration and a plot of particle number versus concentration is compatible with a micelle-like phenomenon; from this curve the critical concentration for the formation of particles is estimated to be approximately one mole of T(is) to 120 moles of lecithin. These data suggest that the membrane-penetrating peptide T(is) is being incorporated, monomerically and multimerically, within the hydrocarbon phase of lecithin bilayers. From these data it can be calculated that each intramembranous particle contains between 10 and 20 T(is) monomers. The peptide portion of each particle, therefore, has a molecular weight of 45,000-85,000. An exact analogy cannot be drawn at this time between the in vivo structure of erythrocyte intramembranous particles and the reconstituted particles described here, although an argument has been constructed to support this possibility. What is clear is that the reconstituted system promises to be useful for further examination of protein-lipid interactions in membranes.

Amino Acid Sequence↗