Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0027960 (mole)
 21,279 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To determine the effect of cholesterol and lipid packing on the solubility of membrane proteins in bilayers, cytochrome b5 incorporation into phosphatidylcholine (PC) liposomes was determined as a function of bilayer curvature (SUVs versus LUVs), fatty acyl chain composition, and cholesterol content. The equilibrium affinity constant for the formation of a 1:1 b5/PC complex, Kp, and the number of PC's per "site" at saturation, n, were determined from binding isotherms, which were obtained by measuring the increase in intrinsic tryptophan fluorescence. With LUVs, n was also determined directly by gel filtration. The following results were obtained: (1) Both Kp and the saturating level of b5 binding, s (n-1), are significantly greater for SUVs than for LUVs. In LUVs, a binding site must consist of several surrounding lipid layers. (2) Cholesterol reduces Kp and s by factors that range from 1 to > 100. Binding inhibition is highly sensitive to the liposome size and to the fatty acyl composition of the PC; the latter correlates with the condensing effects on PC: C1satC2mono > C1satC2di approximately natural mixtures > C1unsat-C2unsat. (3) With POPC LUVs, the binding inhibition was 3.6-, 1.4- and 17-fold within the ranges of 0-20, 20-33, and 33-50 mole percent cholesterol, respectively. (4) The equilibrium binding constant to SUVs is greater for liposomes that are prepared from natural PC mixtures than for vesicles of a single synthetic phospholipid. The reductions in b5 binding correlate with reductions in bilayer free volume, which were calculated from monolayer studies of the lipid mixtures. The sensitivity of liposome saturability to bilayer curvature, fatty acyl chain composition, and cholesterol content may account for the disparate results among previous studies of cholesterol-protein interactions. A more significant implication is that in biological membranes with high levels of cholesterol, subtle variations in the fatty acyl chain composition could substantially affect the solubility and physical states of integral membrane proteins.
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PMID:Effect of cholesterol, fatty acyl chain composition, and bilayer curvature on the interaction of cytochrome b5 with liposomes of phosphatidylcholines. 789 81

The hairpin-duplex equilibria of the dodecamer d-AAGCTTAAGCTT and interaction of the duplex form with a pentapeptide, KGWGK, has been studied. UV thermal transitions are monophasic at low salt but biphasic at higher salt concentrations. At 10(-5) M or less oligomer concentration biphasic melting curves persist till 900 mM NaCl. The d(Tm)/d log(Na+) for the duplex form is 12 degrees C and for the hairpin is 18 degrees C. The delta H and delta S values for duplex formation are low (-25 K cal/mole and -59 Cal/mole respectively). KGWGK binds to the duplex form with a binding constant K = 3.4 x 10(5)M-1 measured from fluorescence quenching of tryptophan. These unusual results are markedly different from that reported for d-AGATCTAGATCT (Biochemistry 31, 6241-6245) and are discussed in terms of sequence dependence of loop folding and cruciform extrusion pathway of hairpin formation.
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PMID:Hairpin formation in d-AAGCTTAAGCTT under high salt conditions shows unusual properties. 794 59

The G protein alpha-subunit G alpha 2 is essential to the developmental program of Dictyostelium. G alpha 2 is transiently phosphorylated on a serine residue(s) following stimulation with extracellular cAMP (Gundersen, R. E., and Devreotes, P.N. (1990) Science 248, 591-593). To aid in defining the function of alpha-subunit phosphorylation, we identified the site of G alpha 2 phosphorylation. Comparison of the isoelectric points (pI) of the phosphorylated and nonphosphorylated forms indicated that a single mole of phosphate is added to G alpha 2. Cleavage at tryptophan residues and immunoprecipitation with a specific peptide antibody localized the phosphorylated serine in the N-terminal 119 residues. Analysis of a series of G alpha 1 and G alpha 2 chimeras further confined the site between amino acids 33 and 215. Site-directed mutagenesis of serines between amino acids 33 and 119 produced two mutants that were not phosphorylated, S45A and S113A. Ser113 was identified as the site by sequential Edman degradation of 32P-radiolabeled G alpha 2 digested with endoproteinase Glu-C. We have expressed the G alpha 2 mutants S113A, S113I, S113T, and S113D in a G alpha 2 null cell line to examine the function of phosphorylation.
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PMID:Serine 113 is the site of receptor-mediated phosphorylation of the Dictyostelium G protein alpha-subunit G alpha 2. 806 9

The pH titration of nine amino acids (glycine, proline, valine, serine, glutamine, tryptophan, arginine, histidine and aspartic acid) in presence of urea in the concentration range 1-8 mole dm-3 has been performed. The results support suppression of the first dissociation constant (K1) of the amino acids and acceptance of H+ ions by the amide forming uronium ion (UH+). The second dissociation constant (K2) of the amino acids is affected relatively weakly by urea. Quantitative evaluation of different species existing in solution and the degree of dissociation of the acids as well as the degree of binding of H+ ion to the amide have been made. It has been found that the polarity of the aqueous-urea medium does not straight forwardly correlate with the altered pK1 of the amino acids. Urea can also affect the pH-titration behaviour of gelatin with an increase of the intrinsic pK of the acidic groups of the protein by 0.45 unit.
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PMID:Interaction of amino acids and gelatin with urea. 814 76

Quantitative studies of the binding of protein kinase C (PKC) to lipid cofactors were performed by monitoring resonance energy transfer with time-resolved fluorescence techniques. For that purpose, diacylglycerol (DG), phosphatidylinositol 4,5-biphosphate (PIP2), phosphatidylinositol 4-phosphate (PIP), phosphatidylinositol (PI), phosphatidylcholine (PC), and phosphatidylserine (PS) were labeled with a pyrenyl decanoyl moiety at the sn-2 position of the lipid glycerol. These labeled lipids proved excellent energy acceptors of light-excited tryptophan residues in PKC. The quenching efficiency of the tryptophan fluorescence was determined as function of lipid probe concentration in mixed micelles consisting of poly(oxyethylene)-9-lauryl ether, PS, and various mole fractions of probe lipid. The experimental conditions and method of data analysis allowed the estimation of binding constants of single or multiple pyrene lipids to PKC. The affinity of PKC for inositide lipids increases in the order PI < PIP < PIP2. The affinity of PKC for PIP and PIP2 is higher than that for DG. Determination of PKC activity in the presence of labeled lipids and PS showed that only PIP2 and DG activate PKC. Double-labeling experiments suggest that PIP2 and DG are not able to bind simultaneously to PKC, indicating a reciprocal binding relationship of both cofactors. The results support the notion that, besides DG, PIP2 can be a primary activator of PKC.
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PMID:Quantitation of the interaction of protein kinase C with diacylglycerol and phosphoinositides by time-resolved detection of resonance energy transfer. 824 Nov 87

The clotting enzyme (Stenoxobin), from the venom of Lachesis muta stenophyrs, was purified by gel chromatography on Bio-gel P-100 followed by agmatine CH-Sepharose-4B and FPLC on Mono Q column. By SDS polyacrylamide gel electrophoresis the mol. wt was found to be 37,000. The enzyme is a glycoprotein with 1.6 moles of sialic acid per mole of protein and has an average content of 7.0% of neutral carbohydrates. The clotting and esterolytic (BAEE) activities were 843 NIH units/mg and 60.1 +/- 1.2 OD225 ml/min/mg, respectively, and could not be inhibited by heparin or hirudin. Amino acid analysis revealed a low content of tryptophan and a high content of acid residues. Stenoxobin acts upon human fibrinogen by releasing consecutively fibrinopeptides A and B from the alpha- and beta-chains of fibrinogen.
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PMID:A thrombin-like enzyme from bushmaster (Lachesis muta stenophyrs) venom. 831 Apr 44

Familial dysalbuminemic hyperthyroxinemia (FDH) is an autosomal dominant syndrome in which clinically euthyroid patients have elevated total thyroxine levels. These high serum thyroxine levels are traceable to altered binding of thyroxine to the patient's albumin. Albumin from FDH patients and normal volunteers have been purified. Reverse-phase and ion-exchange high performance liquid chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis on the FDH-human serum albumin (HSA) samples show a single band that comigrates with normal HSA. In both protein solutions the intrinsic fluorescence, upon 280 nm excitation, is predominantly due to the single tryptophan residue. The quantum yield of this intrinsic fluorescence in the FDH-HSA solutions is, however, reduced relative to that of HSA. Furthermore, the "average" lifetime value of the tryptophan emission in the FDH-HSA sample is less than that of normal HSA, consistent with its reduced quantum yield. The binding of thyroxine to both albumins effectively quenches the tryptophan emission probably via a nonradiative energy transfer mechanism. Time-resolved data suggest that the albumin from the dysalbuminemic patients is actually an approximately equimolar mixture of normal HSA and FDH-HSA indicative of heterologous expression. Quenching of the intrinsic HSA and FDH-HSA fluorescence by serial additions of thyroxine showed enhanced quenching of FDH-HSA relative to HSA at any T4 to albumin mole ratio, therefore supporting earlier reports of increased thyroxine affinity to FDH-HSA.
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PMID:Fluorescence investigations of albumin from patients with familial dysalbuminemic hyperthyroxinemia. 847 73

Beta-globulin, a single polypeptide chain of molecular weight 15,000 +/- 1,000, undergoes denaturation in alkaline pH (7.0-13.0), thereby affecting the hydrodynamic properties of the protein, viz. a decrease in sedimentation coefficient from a value of 2.0s to 1.4s at pH 11.3, an increase in reduced viscosity from 0.042 dl/g to 0.158 dl/g at pH 12.6 and a decrease in partial specific volume resulting in a volume change of 6.3 +/- 1.0 ml/mole residue at pH 11.7. The perturbation of tryptophanyl residues and ionization of tyrosyl residues are preceded by alteration in conformational status of the protein. The fluorescence emission measurements indicate initial unfolding of the protein molecule which exposes the tryptophan and tyrosyl residues to the solvent. The tyrosyl phenolic group ionization is anomalous having a pKint value of 11.2. The reduced viscosity value reaches a plateau region at pH 12.5.
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PMID:Structural stability of beta-globulin, the low molecular weight protein fraction from sesame seed (Sesamum indicum L.) in alkaline solution. 850 22

The fluorescence of a membrane-bound tryptophan derivative (tryptophan octyl ester, TOE) has been examined as a model for tryptophan fluorescence from proteins in membrane environments. The depth-dependent fluorescence quenching of TOE by brominated lipids was found to proceed via a dynamic mechanism with vertical fluctuations playing a central role in the process. The activation energy for the quenching was estimated to be 1.3 kcal/mole. The data were analyzed using the distribution analysis (DA) method, which extends the conventional parallax method to account more realistically for the transbilayer distributions of both probe and quencher and for possible variations in the probe's accessibility. DA provides a better fit than the parallax method to data collected with TOE in membranes formed of lipids brominated at either the 4,5, the 6,7, the 9,10, or the 11,12 positions of the sn-2 acyl chain. DA yields information on the fluorophore's most probable depth in the membrane, its conformational heterogeneity, and its accessibility to the lipid phase. Previously reported data on cytochrome b5 and melittin were reanalyzed together with data obtained with TOE. This new analysis demonstrates conformational heterogeneity in melittin and provides estimates of the freedom of motion and exposure to the lipid phase of membrane-embedded tryptophans of cytochrome b5.
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PMID:Fluorescence of membrane-bound tryptophan octyl ester: a model for studying intrinsic fluorescence of protein-membrane interactions. 1152 96

Binding of a tetra peptide, lysyl tryptophenyl glycyl lysine O-ter butyl ester (KWGK) with duplex forms of G, C containing polynucleotides, Poly [d(G-C)], Poly [d(G-5M C)], Poly (dG), Poly (dC) and E.coli DNA were studied under low salt conditions using UV absorption, fluorescence, and circular dichroic (C.D) spectroscopy. On addition of the peptide (upto a P/N mole ratio of 0.5), the Poly [d(G-5M C)] under low salt (1 mM Na Cl) conditions, was converted from Z to B-form as shown by the inversion of C.D spectra. The two binding constants (K1 and K2) were determined from fluorescence spectroscopy of which K2 estimates the intercalation of the tryptophenyl side chain between the base pairs of DNA and K1 estimates the electrostatic interactions between the lysyl side chains and phosphate groups. The strength of intercalation is: Z-form of Poly [d(G-5M C)] >> B form of Poly [d(G-5M C)] >> E.Coli DNA > Poly (dG).Poly (dC). This means that peptide seems to have strong preference for Z compared to B-form and for alternating over non-alternating G, C Sequences. This suggests that tryptophan intercalation may act as a discriminating factor in recognizing Z and B-forms and may have a potential role in Protein-Nucleic acid interactions that are important for transcription.
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PMID:Tryptophan intercalation in G, C containing polynucleotides: Z to B conversion of poly [d(G-5M C)] in low salt induced by a tetra peptide. 887 59


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