Search PubMed⌕ Search

Biomedical subjects

J B Swaney

Publications and source records attributed to J B Swaney.

At least 37 records · Page 2Linked to original sources
PubMed1984

Hybrid association between human apolipoproteins A-I and A-II in aqueous solution and in phospholipid recombinants.

The formation of hybrid association products between apolipoprotein A-I and apolipoprotein A-II from human high-density lipoprotein was investigated in solutions of these apolipoprotein and in recombinant particles with dimyristoylphosphatidylcholine (DMPC). It was found that these two proteins interact in solution to form hybrid association products, but not to a marked degree. When these two proteins were incubated together with DMPC, it was likewise found that there was little tendency to reside on the same particle, as judged from the absence of hybrid oligomers by chemical cross-linking. By a modified immunoelectrophoretic method it was found that only about 15% of the A-II and 10% of the A-I were precipitated by the heterologous antiserum; from this it is concluded that 80-90% of these proteins do not form hybrid recombinants with the other protein. These results suggest that in the delipidated state, as well as in discoidal recombinants, there do not exist strong protein-protein interactions between A-I and A-II. This implies that even in the high-density lipoprotein, where both proteins coexist in the same particle, the A-II does not stabilize the molecular structure through interactions with A-I, and its role in this molecule remains obscure.

Apolipoprotein A-I↗
PubMed1983

Reconstitution of apolipoprotein A-I from human high density lipoprotein with bovine brain sphingomyelin.

Reconstitution of apolipoprotein A-I was found to occur readily with bovine brain sphingomyelin (BBSM), with a maximum rate occurring at a temperature of 28 degrees C, a temperature approximating the phase transition temperature for this naturally occurring phospholipid. At BBSM:A-I weight ratios of 7.5:1 or less, a single recombinant product was observed which contained three A-I molecules per particle, which had a BBSM:A-I molar ratio of 360 to 1 and which appeared in the electron microscope as a discoidal complex with a thickness of 68 A and a diameter of 217 A. By these criteria, as well as by gel filtration, this product appears very similar to that obtained by recombination of A-I with phosphatidylcholine at elevated ratios of phospholipid/protein. No evidence was found for the existence of any BBSM:A-I complexes comparable to the smaller lecithin:A-I complex containing 200-250 mol of phospholipid and two A-I molecules per complex which has been previously reported. At BBSM:A-I ratios of 15:1 (w/w), a new type of complex was observed which was discoidal by electron microscopy but possessed a larger diameter (390 A) and higher phospholipid:protein molar ratio (535:1) than has been observed previously for recombinant complexes. The BBSM:A-I complexes were found to be significantly more resistant to denaturation by guanidine hydrochloride than the dimyristoyl phosphatidylcholine:A-I recombinant complexes. It is concluded that the mechanisms of interaction between apolipoprotein A-I and either bovine brain sphingomyelin or phosphatidylcholines are similar, but that the nature of the protein-lipid interactions with BBSM are such as to produce larger and more stable complexes than are observed with the phosphatidylcholines.

Animals↗
PubMed1983

Selective proteolytic digestion as a method for the modification of human HDL3 structure.

Trypsin digestion of human high density lipoprotein (d 1.125-1.21 g/ml) on which the lysine residues have been masked with the reversible blocking group, 2,3,4,5-tetrahydrophthallic anhydride (THPA), was found to result in the fragmentation of the apoA-I component, but not the apoA-II component of this lipoprotein particle. Approximately 50-80% of the apoA-I polypeptide was found in a lipid-free fraction, while the residual apoA-I material plus the apoA-II moiety constituted a core particle that contained most of the original lipid. Immunological analysis indicated that such fragmentation did not affect the immunoreactivity of apoA-II, but that all immunoreactivity of apoA-I was lost within the first 30 min of trypsinization. By column chromatography and electron microscopy this core particle appeared identical in size with the untrypsinized THPA-modified HDL(3) material. Size analysis of the core particle peptides suggests that not all of the A-I molecules present on the HDL(3) are trypsinized to the same extent, which indicates possible nonequivalence of these peptide chains. Analysis of the amino acid composition revealed a somewhat greater proportion of hydrophobic residues in the lipid-bound fraction than in the lipid-free fraction. Analysis of tryptophan showed that almost all of this highly hydrophobic residue was found in the lipid-bound fraction; this suggests that lipid binding occurs preferentially in the more hydrophobic domains of the A-I molecule. Incubation of the core particle with intact apoA-I, obtained from either human or bovine HDL, showed that these proteins could be incorporated to regenerate an HDL(3) of selectively altered protein composition, compared to the original lipoprotein. It is concluded that some latitude is allowable in the surface/volume relationship in lipoproteins before reorganization of the particle is required; this might, for example, provide a mechanism whereby the HDL could serve in a storage role for the C apolipoproteins in plasma.-Swaney, J. B. Selective proteolytic digestion as a method for the modification of human HDL(3) structure.

Amino Acids↗
PubMed1980

Thermal dependence of apolipoprotein A-I-phospholipid recombination.

Studies of the recombination of apolipoprotein A-I (apo A-I), the major protein constituent of human high-density lipoprotein, and various synthetic phospholipids, both alone and in mixtures, have been performed. Pure diacyl phospholipids containing homologous fatty acids of the C12, C13, C14, and C15 series, as well as the two positional isomers containing C14 and C16 fatty acids in positions 1 and 2, undergo reaction with the apo A-I protein only near their gel-liquid-crystalline transition temperatures; the degree of reactivity of these phospholipids toward recombination was observed to decrease as the transition temperature increased. The presence of lysolecithin in the incubation mixtures at proportions of 5 mol/mol of protein or lower was not found to have a significant effect on the rate of recombination. Binary mixtures of dimyristoylphosphatidylcholine and dipalmitoylphosphatidylcholine at various proportions react maximally with apo A-I at the onset of the phase transition, as judged by comparison with published phase diagrams; in this case also, the rate of recombination was observed to decline for mixtures with higher phase transition temperatures. These results are interpreted in terms of protein insertion at lattice defects arising from the presence of phospholipid clusters undergoing the phase transition; these clusters are derived from the cooperative and simultaneous melting of a number of molecules, the cooperativity being dependent upon the nature of the phospholipid. It is postulated that phospholipids which melt in a more highly cooperative fashion are more capable of interacting with the apolipoproteins since these phospholipids will possess larger lattice defects during the phase transition.

Apolipoprotein A-I↗
PubMed1980

Properties of lipid-apolipoprotein association products. Complexes of human apo AI and binary phospholipid mixtures.

The products resulting from the association of human apo A-I with binary phospholipid mixtures of dimyristoyl phosphatidylcholine (DMPC) and either dipalmitoyl (DPPC) or distearoyl (DSPC) phosphatidylcholine have been isolated and characterized. Effective lipid . protein complex formation was found to occur at the onset temperature for melting of the gel state, and equal incorporation of both lipid components of the binary mixture was observed. Two sizes of products were obtained, one containing 2 A-I molecules per complex and the other containing 3; the proportions of these two products depended upon the initial phospholipid/protein ratio employed. these two product species were found to be resolvable by density gradient centrifugation as well as gel filtration, reflecting substantial differences in composition as well as size. The ratio of DMPC to DPPC or DSPC was the same in the isolated complexes as in the initial mixture, suggesting that th protein does not associate preferentially with the fluid phase lipid, but with lipid domains in which the components are randomly distributed. Electron microscopy of recombinant particles containing a 2:1 ratio (w/w) of DSPC to DMPC revealed stacks of discs whose thickness was proportionately greater than for discs containing DMPC alone. Also of significance was the finding that recombinant discs containing 3 A-I molecules possessed a diameter approximately 1.5 times larger than recombinant discs containing 2 A-I molecules. These data are consistent with the model for the recombinant particles described by Tall et al. (Tall, A.R., Small, D.M., Deckelbau, R.J., and Shipley, G.G. (1977) J. Biol. Chem. 252; 4701-4711), in which the phospholipid is found as a circular bilayer, the thickness of which is dependent upon the length of the acyl chain, and around which the protein is distributed as an annulus.

Apolipoprotein A-I↗
PubMed1980

Mechanisms of protein-lipid interaction. Association of apolipoproteins A-I and A-II with binary phospholipid mixtures.

Studies of the recombination of apo A-I and apo A-II the major protein components of human high density lipoprotein, with binary mixtures of dimyristoyl phosphatidylcholine (DMPC) and distearoyl phosphatidyl-choline (DSPC) were performed. Recombination was observed to occur only near the lower bound temperature of the phase transition for each mixture. Similar experiments using binary mixtures of DMPC and 1-palmitoyl-2-oleoyl phosphatidylcholine (POPC) demonstrated that recombination occurs in a temperature range which is believed to approximate the lower bound of the phase transition for this mixture as well. The reactivity of both types of mixtures toward recombination with apolipoproteins was found to decrease with decreasing proportions of DMPC, even though the effect of DMPC was to decrease the transition temperature in DMPC/POPC mixtures and to increase the transition temperature in DMPC/DSPC mixtures. A mechanism for insertion of apolipoproteins into lipid bilayers is proposed in which the protein gains entry to the interior of the bilayer through a defect resulting from equilibrium fluctuations of state at the onset temperature of acyl chain melting.

Apolipoproteins↗
PubMed1980

Characterization of the high-density lipoprotein and its major apoprotein from human, canine, bovine and chicken plasma.

The high-density lipoproteins (HDL) from canine, bovine, and chicken plasma have been shown to contain almost exclusively the apolipoprotein A-I, while human HDL contains a second major component, the apolipoprotein A-II. Chemical cross-linking demonstrated that dog and chicken HDL contain three apolipoprotein A-I molecules per particle, while bovine HDL contain approximately six apolipoprotein A-I molecules per particle. By this method, the amount of protein in human HDL2 (d = 1.063-1.12) was found to be approximately 120 000 g/mol, while for human HDL3 (d = 1.12-1.21) a value of approximately 90 000 g/mol was obtained, suggesting that the protein complement of HDL2 and HDL3 differ by only one apolipoprotein A-I chain per particle. Comparison of the apolipoprotein A-I from various animal species indicated that the canine and human apolipoprotein A-I proteins were the most similar by fluorescence, self-association properties, and immunoreactivity. Cross-linking of chicken and bovine apolipoprotein A-I yielded patterns distinctly different from that obtained with the human or canine counterpart. It is concluded that the quaternary structure of the various species of HDL is not directly correlated with the degree of self-association found for the protein constituents.

Amino Acids↗
PubMed1979

Effects of estrogen administration on the lipoproteins and apoproteins of the chicken.

The plasma lipoproteins of estrogen-treated and untreated sexually immature hens have been compared with respect to their concentration in plasma, protein and lipid composition, particle size, and and apoprotein composition. Administration of diethylstilbestrol resulted in a 400-fold rise in the concentration of very low density lipoprotein (VLDL), a 70-fold rise in low density lipoprotein (LDL), and a marked reduction in high density lipoprotein (HDL) protein. It also resulted in the production of LDL and HDL which were enriched in triacylglycerol, while the proportion of cholesterol in all three lipoprotein fractions decreased. In contrast to the lipoproteins from untreated birds, lipoproteins of density less than 1.06 g/ml from estrogen-treated birds were not clearly separable into discrete VLDL and LDL fractions, but appeared to be a single ultracentrifugal class. The apoprotein composition of VLDL and LDL from untreated birds differed from each other; however, the apoprotein patterns of VLDL and LDL from estrogen-treated birds were indistinguishable: both contained a large amount of low molecular weight protein in addition to the high molecular weight component that predominates in the untreated state. The apoprotein composition of HDL was also markedly altered by estrogen administration: the 28,000 mol. wt. protein (apo A-I) decreased in amount from 65% to less than 5% of the total, while a low molecular weight (Mr = 14,000) protein and as yet poorly defined high molecular weight components became predominant. These observations indicate that the hyperlipidemia induced by estrogen administration is accompanied by marked alterations, both qualitative and quantitative, in the plasma lipoproteins.

Animals↗
PubMed1978

Cross-linking studies of the self-association properties of apo-A-I and apo-A-II from human high density lipoprotein.

The state of self-association of the apoprotein components of human high density lipoprotein have been studied by use of the cross-linking reagent dimethyl-suberimidate. Analusis of the cross-linked products was carried out by soduim dodecyl sulfate-gel electrophoresis and by agarose column chromatography in 6 M guanidine hydrochloride. Apo-A-I was found to exist as a monomer at low concentration, but associates to tetrameric and pentameric forms at concentrations of 0.5 mg/ml or higher. The self-association was found to be ionic strength-dependent, with association promoted by the presence of salt. Apo-A-II was also found to associate, but the major oligomeric form observed was dimeric (Mr = 34,000), and the association was less dependent on ionic strength than for apo-A-I. Cross-linking in the presence of various concentrations of guanidine hydrochloride showed that apo-A-II self-association persisted at higher concentrations of the denaturant than for apo-A-I. Studies of the effect of temperature demonstrated that the self-association of both proteins was diminished at temperatures above 30 degrees C. Recombination of apo-A-II with phospholipid resulted in the formation of particles which yielded primarily trimers upon cross-linking. This suggests that phospholipid binding causes major reorganization of the self-associated forms of apo-A-II.

Amino Acids↗
PubMed1977

Characterization of the apolipoproteins of rat plasma lipoproteins.

Purified fractions of three major rat high-density lipoproteins (HDL) and one rat very low-density lipoprotein (VLDL) were isolated by Sephadex gel chromatography or preparative sodium dodecyl sulfate gel electrophoresis. These proteins were characterized by amino acid analysis, end-group analysis, molecular-weight determination, polyacrylamide gel electrophoresis, and circular dichroism. One of these rat proteins, of molecular weight 27 000, appears to be homologous with the human A-I protein. However, rat HDL possesses two additional major components not reported in human HDL - an arginine-rich protein of molecular weight 35 000 and a protein of molecular weight 46 000. The arginine-rich protein of the rat is similar in size and amino acid analysis to the arginine-rich protein reported in human VLDL. A major component of rat VLDL of 35 000 molecular weight appears similar or identical to the arginine-rich protein in rat HDL by every criterion employed for their characterization.

Amino Acid Sequence↗
PubMed1977

Analysis of rat serum apolipoproteins by isoelectric focusing. II. Studies on the low molecular weight subunits.

The low molecular weight proteins of rat apo HDL and apo VLDL have been isolated and analyzed by the technique of isoelectric focusing. Sephadex fractions from apo HDL (HS-3) and apo VLDL (VS-3) that contain these proteins reveal three major bands with apparent isoelectric points of pH 4.50, 4.67, and 4.74, as well as three minor bands at pH 4.43, 4.57, and 4.61. In addition, apo HDL has a major band at pI of 4.83. DEAE-Cellulose chromatography was used to prepare purified fractions of these components that were characterized by N-terminal analyses and molecular weight determinantions by SDS gel electrophoresis. The major low molecular weight components of apo HDL were focused on a slab gel and the bands were identified as A-II (pI 4.83), C-II (pI 4.74), C-III-0 (pI 4.67), and C-III-3 (pI 4.50). Neuraminidase treatment of apo HDL, followed by isoelectric focusing, suggested that the other bands, which have not previously been reported, may be additional forms of the C-III protein, differing only in their content of sialic acid.

Animals↗
PubMed1977

Analysis of rat serum apolipoproteins by isoelectric focusing. I. Studies on the middle molecular weight subunits.

Analytical isoelectric focusing (IEF) has been applied to the study of the apolipoprotein components of rat serum high density and very low density lipoproteins. The apolipoproteins were separated on 7.5% polyacrylamide gels containing 6.8% urea, with a pH gradient of 4-6. The middle molecular weight range apolipoproteins were identified on IEF gels by the use of apolipoproteins purified by electrophoresis on gels containing sodium dodecyl sulfate (SDS). The A-1 protein focused as 4 to 5 bands from pH 5.46 to 5.82; the A-IV protein and the arginine-rich protein each focused as 4 to 6 bands from pH 5.31 to 5.46. The low molecular weight proteins focused from pH. 4.43 to 4.83 and are the subject of a separate communication. Comparisons of the IEF method with SDS gel electrophoresis, polyacrylamide gel electrophoresis in urea, and Sephadex chromatography are also reported. Additional studies were also carried out that tend to rule out carbamylation or incomplete unfolding of the proteins in the presence of urea as the causes of the observed heterogeneity.

Amino Acids↗