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H Edelhoch

Publications and source records attributed to H Edelhoch.

At least 55 records · Page 3Linked to original sources
PubMed1978

Self-association of glucagon as measured by the optical properties of rhodamine 6G.

The fluorescence of rhodamine 6G is completely quenched in glucagon solutions in 0.6 M K2HOP4 at pH 10.6. The absorption of rhodamine 6G is red-shifted by the same reaction. A single rhodamine 6G molecule appears to be bound to a hydrophobic patch in the center of the trimer of glucagon. Since the glucagon monomer has almost no organized structure this site exists only in the associated trimer form of glucagon. The self-association of glucagon to the trimer has been determined from the variation in rhodamine 6G fluorescence and absorption measured over a 60-fold range of dye concentration. The self-association constant agrees with values determined by other methods in the absence of dye. The binding isotherms of rhodamine 6G to glucagon shift with glucagon concentration and exhibit negative cooperativity.

Anilino Naphthalenesulfonates↗
PubMed1978

Protein-lipid interactions and the role of water.

The rigidity of the three-dimensional structure of a native protein is dependent on the network of hydrogen-bonded groups which provide the scaffolding for the other interactions. The structure is stabilized by the hydrophobic interactions of the nonpolar side chains. The latter are formed by the very unfavorable entropy change that occurs in water but not in less-polar solvents. It is unlikely that any solvent other than water can produce the same folding of a polypeptide chain to form the active native structure. Water plays a unique role, since it alone is responsible for the heat capacity changes observed when nonpolar groups are transferred from an aqueous to a nonaqueous environment, as exists in the interior of a protein. The need to juxtapose like groups and to avoid making contact among unlike groups imposes severe restrictions on the binding of small or large molecules to proteins. Consequently there must be proper pairing of polarities as well as close fitting of ligands for strong binding to occur. This is clearly evident from the x-ray studies of proteins containing subunits or prosthetic groups. The thermodynamic parameters observed in the most complex protein reactions--i.e., self-assembly systems--resemble rather well those observed in micelle association reactions or even in the solution of nonpolar gases in water. This interaction--hydrophobic--can be looked upon as the controlling reaction which stabilized the organized structures of most cellular entities aside from nucleic acids--i.e., membranes and organelles.

Butanes↗
PubMed1977

Structure and stability of human thyroxine-binding globulin.

The secondary and tertiary structure of human plasma thyroxine-binding globulin (TBG) was investigated by circular dichroism and fluorescence properties. The relaxation time of TBG indicated that it is a compact, symmetric molecule. It was calculated from the far ultraviolet CD spectrum that about one-half of the peptide groups are equally distributed in alpha helical and beta structures. In the near ultraviolet, the CD spectrum of TBG was modified when thyroxine was bound. TBG was stable at temperatures below 50 degrees at pH 9 and below 35 degrees at pH 10.5. Below pH 5 tryptophanyl fluorescence revealed a molecular transition which followed first order kinetics. The transition resulted in an irreversible loss of binding of the hormone. Acidification to pH 3.4 produced only a minor change in the CD spectrum, in which some of the alpha helical peptides were converted to beta structure.

Circular Dichroism↗
PubMed1977

Interaction of apoA-II from human high density lipoprotein with lysolecithin.

The effects of lysolecithin and hexadecyltrimethylammonium bromide on the structure and stability of apoA-II from human high density lipoprotein have been evalued by circular dichroism and fluorescence measurements. There is a profound enhancement in the stability of apoA-II to guanidinium hydrochloride denaturation when it forms a phospholipid complex with lysolecithin micelles. This complex is not only resistant to guanidinium hydrochloride denaturation, but it can be formed in a 6 M solution of this denaturant. The behavior of apoA-II in the native human high density protein is much closer to that of the lysolecithin apoA-II complex than to that of the free apoA-II.

Apoproteins↗
PubMed1977

Stability and subunit structure of human alpha2-macroglobulin.

The molecular weights of alpha2-macroglobulin and its non-covalent subunits have been determined by equilibrium centrifugation. The secondary structure of the native and the thermally denatured molecules has been analyzed by circular dichroic measurements. In contrast to most proteins the thermally denatured form contains a slightly more highly organized polypeptide chain than the native form. The relaxation time of the native protein, as determined by fluorescence polarization measurements, indicates that alpha2-macroglobulin is composed of domains smaller than that of the two subunits. The transitions in acid, alkali, and at high temperatures have been explored in order to establish the pH and thermal range of stability of alpha-macroglobin.

Drug Stability↗
PubMed1977

Thermodynamics of the self-association of glucagon.

In water, glucagon exists in an equilibrium between a trimer in which more than half of the peptide groups are in an alpha-helical configuration and a monomer which has a random coil configuration with few alpha-helical residues. The thermodynamics of this self-association have been evaluated by studying the temperature- and concentration-dependence of the mean residue ellipticity at 220 nm. The enthalpy and entropy changes of association were negative at all temperatures between 5 degrees and 50 degrees and had large negative temperature dependencies. Usually an association that involves nonpolar groups is considered to be driven by a positive entropy term. Such an explanation is not tenable in the case of glucagon. However, if the effects of nonpolar groups on the coil-to-helix transition of a polypeptide are included into the thermodynamic considerations of hydrophobic interactions, then the negative parameters observed for glucagon association can be readily understood. The hydrophobic interaction is therefore not necessarily controlled by the entropy change because, if there are significant conformational changes, the reaction may be controlled by the enthalpy change. Consequently, the more important parameter characteristic of all hydrophobic reactions is the heat capacity change.

Glucagon↗
PubMed1975

Ligand-induced self-association of human luteinizing hormone. Negative cooperativity in the binding of 8-anilino-1-naphthalensulfonate.

The self-association of human luteinizing hormone (hLH) is enhanced in the presence of 8-anilino-1-naphthalenesulfonate (ANS). Sedimentation equilibrium measurements indicate that the hormone exists primarily as a dimer in the presence of excess ANS. It is shown that, for a self-associating protein system in which monomer and dimer have different affinities and/or capacities for ligand, both the shape and the position of the binding curve depend on protein concentration. Gel filtration and fluorescence measurements indicate that the hLH dimer has a single high affintiy site (K greater than 10(6) M-1) for ANS while binding to the monomer is too weak to be observed. This leads to negative cooperativity in the binding and to a shift of the binding curve to lower free ligand concentration with increasing concentration of the hormone.

Anilino Naphthalenesulfonates↗
PubMed1975

Ligand-induced self-association of human chorionic gonadotropin. Positive cooperativity in the binding of 8-anilino-1-naphthalenesulfonate.

Human chorionic gonadotropin (hCG) self-associates to form higher molecular weight species in the presence of the fluorescence probe 8-anilino-1-naphthalenesulfonate (ANS). Sedimentation equilibrium and fluorescence titration data have been analyzed in terms of a monomer-dimer-tetramer model in which the various oligomers have different affinities and/or capacities for the ligand. The results indicate that the ligand affinities are in the order tetramer greater than dimer greater than monomer whereas the numbers of ligand binding sites per mole of hCH are in the reverse order. Consequently, addition of ANS first shifts the equilibrium from monomer to tetramer and gives rise to positive cooperativity in the titration curves. At sufficiently high ANS concentration (approximately 0.5 mM), the equilibrium shifts back to the dimer because of its greater binding capacity. This is manifested by a second phase in the titration curve and a decrease in the polarization of ANS fluorescence. The results are discussed in terms of the general problem of ligand controlled protein association and are contrasted to results reported to the previous paper for the homolgous protein, human luteinizing hormone.

Anilino Naphthalenesulfonates↗
PubMed1975

Relationship between iodination and the polypeptide chain composition of thyroglobulin.

Thyroglobulin was isolated from thyroid glands of normal guinea pigs and from animals treated with thiouracil. These preparations were fractionated by isopyknic centrifugation in RbCl into proteins of varying iodine content. When the disulfide bonds of these protein fractions were reduced and analyzed by polyacrylamide gel electrophoresis in Na dodecyl-SO4, t hree species were observed with molecular weights of 295,000 (A), 210,000 (B), and 110,000 (C). Species A comprised 80% of the protein in thyroglobulin of 0.04% iodine and 13% in thyroglobulin of 0.68% iodine content. Species C showed the opposite relationship, comprising 10% of the low and 70% of the high iodine thyroglobulin. Species B was relatively independent of the iodine content and represented approximately 20% of the protein. Iodine analysis of these proteins showed species A to be lowest and species C highest. It appears that the subunit composition of thyroglobulin depends on the degree of iodination and that species A should be the only one present in the absence of iodination.

Animals↗
PubMed1975

Elementary chain composition of guinea pig thyroglobulin.

Thyroglobulin obtained from guinea pigs was examined by Na dodecyl-SO4-polyacrylamide gel electrophoresis after reduction and alkylation. In contrast to thyroglobulin from other mammalian sources, only three groups of polypeptide chains accounted for 95% or more of the protein. Determinations of the molecular weights of these purified proteins by equilibrium centrifugation in 6 M guanidine HCl gave values of 295,000 (species A), 210,000 (species B), and 110,000 (species C). Molecular weights determined by gel filtration in 6 M guanidine HCl gave similar results. Due to the large size of the polypeptides, satisfactory molecular weights could not be obtained from Na dodecyl-SO4-polyacrylamide gel electrophoresis. Amino acid analysis of the three species was similar to that of whole thyroglobulin. Only slightly higher level of lysine and histidine and a lower level of glutamic acid were seen in species C. The iodine contents were found to range from 0.07 to 0.12 to 0.20% for species A, B, and C, respectively.

Amino Acids↗
PubMed1975

The interaction of apoA-I from human high density lipoprotein with lysolecithin.

The binding of apoA-I to lysolecithin has been studied by fluorescence and circular dichroism. The influence of the conformation of apoA-I on its interaction with lysolecithin has also been evaluated. ApoA-I is bound to lysolecithin with an association greater than 10(7) whether apoA-I is native or highly unfolded in 1.8 M guanidinium hydrochloride. The association of apoA-I with lysolecithin results in an increase in secondary structure. A 25-residue fragment of apoA-I binds to lysolecithin equally strongly as the native molecule.

Apoproteins↗
PubMed1975

The self-association of the reduced ApoA-II apoprotein from the human high density lipoprotein complex.

The molecular properties of the single linear chain form of human apoA-II, i.e., Cm apoA-II, have been evaluated by circular dichroism, polarization of fluorescence, difference absorption, and sedimentation equilibrium. The self-association of Cm apoA-II to a dimer resembles closely that of apoA-II though the free energy change is somewhat smaller. The dimerization of Cm apoA-II is accompanied by major changes in secondary and tertiary structure. The apoA-II molecule, therefore, represents a molecular association where the intramolecular structure is strongly dependent on the quaternary structure.

Apoproteins↗
PubMed1975

The molecular behavior of apoA-I in human high density lipoproteins.

The properties of apoA-I in human high density lipoprotein (1.063 smaller than d smaller than 1.210 gm per ml) have been examined by fluorescence and difference absorption spectroscopy, while the behavior of the total complement of apoproteins has been evaluated by far ultraviolet circular dichroism. Marked increases in stability to temperature, pH, and guanidine hydrochloride were observed for apoA-I in the native particle as compared to the isolated state.

Apoproteins↗
PubMed1975

Negative cooperativity in the binding of thyroxine to human serum prealbumin. Preparation of tritium-labeled 8-anilino-1-naphthalenesulfonic acid.

The binding of thyroxine (T4) and 8-anilino-1-naphthalenesulfonic acid (ANS) to human serum prealbumin was measured by equilibrium dialysis at pH 7.4 in 0.05 M phosphate-0.10 M NaCl at 25 degrees. The data were analyzed for the binding constants based on equations for (1) two independent sites and (2) two identical sites with negative interaction. Evaluation by the independent site model gave the following association constants: for T4 binding, KT1 = 1.0 x 10-8 M-1, KT2 = 9.5 x 10-5 M-1; for ANS binding, KA1 = 9.5 x 10-5 M-1, KA2 = 2.1 x 10-5 M-1. The interactive model gave constants kT = 5.5 x 10-7 M-1 and kA = 5.5 x 10-5 M-1. Interaction factors, alpha, defined such that -RT in alpha is the energy of interaction, were: alpha T = 0.041 AND ALPHA A = 0.62 for T4 and ANS, respectively. The "best fit" values for the number of sites were 2.0 and 1.6 for T4 and ANS, respectively. The binding of T4 to human prealbumin was competitive with ANS, and the binding constants evaluated from competition experiments were in agreement with those found for each ligand when studied separately. On the basis of analysis of X-ray data of human prealbumin (Blake et al.) there appear to be two identical T4 sites. It is therefore evident that the binding of T4 represents a case of negative cooperativity which is presumably due to interaction between ligands.

Anilino Naphthalenesulfonates↗