Biomedical subjects

B de Kruijff

Publications and source records attributed to B de Kruijff.

At least 181 records · Page 10Linked to original sources

PubMed1987

Importance of the tryptophans of gramicidin for its lipid structure modulating activity in lysophosphatidylcholine and phosphatidylethanolamine model membranes. A comparative study employing gramicidin analogs and a synthetic alpha-helical hydrophobic polypeptide.

The importance of the tryptophan residues of gramicidin for the lipid structure modulating activity of this pentadecapeptide was investigated by studying the interaction of gramicidin analogs A, B, C (which have a tryptophan, phenylalanine and tyrosine in position 11, respectively) and tryptophan-N-formylated gramicidin (in which the four tryptophan residues have been formylated) with several phospholipid systems. In addition an alpha-helical model pentadecapeptide (P15) was studied to further test the specificity of the gramicidin-lipid interaction. DSC experiments showed that all the gramicidin analogs produced a significant decrease in the gel to liquid-crystalline transition enthalpy of dipalmitoylphosphatidylcholine. The P15 peptide was much less effective in this respect. In dielaidoylphosphatidylethanolamine the gel----liquid-crystalline transition enthalpy was much less affected by the incorporation of these molecules. In this lipid system tryptophan-N-formylated gramicidin was found to be the most ineffective. 31P-NMR and small angle X-ray diffraction experiments showed that the ability of the peptides to induce bilayer structures in palmitoyllysophosphatidylcholine and HII phase promotion in dielaidoylphosphatidylethanolamine systems follows the order: gramicidin A' (natural mixture) approximately equal to gramicidin A greater than gramicidin B approximately equal to gramicidin C greater than tryptophan-N-formylated gramicidin greater than P15. These results support the hypothesis that the shape of gramicidin and its aggregational behaviour, in which the tryptophan residues play an essential role, are major determinants in the unique lipid structure modulating activity of gramicidin.

Calorimetry, Differential Scanning↗

PubMed1987

Effects of adriamycin on respiratory chain activities in mitochondria from rat liver, rat heart and bovine heart. Evidence for a preferential inhibition of complex III and IV.

The inhibition of respiratory chain activities in rat liver, rat heart and bovine heart mitochondria by the anthracycline antibiotic adriamycin was measured in order to determine the adriamycin-sensitive sites. It appeared that complex III and IV are efficiently affected such that their activities were reduced to 50% of control values at 175 +/- 25 microM adriamycin. Complex I displayed a minor sensitivity to the drug. Of the complex-I-related activities tested, only duroquinone oxidation appeared sensitive (50% inhibition at approx. 450 microM adriamycin). Electron-transfer activities catalyzed by complex II remained essentially unaltered up to high drug concentrations. Of the activities measured for this complex, only duroquinone oxidation was significantly affected. However, the adriamycin concentration required to reduce this activity to 50% exceeded 1 mM. Mitochondria isolated from rat liver, rat heart and bovine heart behaved essentially identical in their response to adriamycin. These data support the conclusion that, in these three mitochondrial systems, the major drug-sensitive sites lie in complex III and IV. Cytochrome c oxidase and succinate oxidase activity in whole mitochondria exhibited a similar sensitivity towards adriamycin, as inner membrane ghosts, suggesting that the drug has direct access to its inner membrane target sites irrespective of the presence of the outer membrane. By measuring NADH and succinate oxidase activities in the presence of exogenously added cytochrome c, it appeared that adriamycin was less inhibitory under these conditions. This suggests that adriamycin competes with cytochrome c for binding to the same site on the inner membrane, presumably cardiolipin.

Animals↗

PubMed1987

Effects of the anti-cancer drug adriamycin on the energy metabolism of rat heart as measured by in vivo 31P-NMR and implications for adriamycin-induced cardiotoxicity.

In vivo 31P-NMR was used to measure the effects of the anti-tumor drug adriamycin on the energy metabolism of rat heart. The exclusive acquisition of NMR signal from cardiac muscle was assured by positioning a solenoidal radio-frequency NMR coil around the heart. Appropriate control experiments verified that 31P-NMR spectra solely originated from this organ. Acute effects occurring shortly after adriamycin administration are expressed in 31P spectra as a dose-dependent decline in the cardiac levels of phosphocreatine, after which stabilization at a new steady-state level occurs. These acute effects of a single dose are complete in 30-60 min and no significant further changes take place within 150 min after drug introduction. Longer-term effects of single high doses and of multiple lower doses were measured up to a week after the initiation of treatment. It seemed that at a total dose of 20 mg/kg, drug-induced interference with cardiac energy metabolism was more pronounced than at the same dose in the acute phase. These 31P-NMR data demonstrate that adriamycin treatment is accompanied by a decrease of the cardiac phosphocreatine/ATP ratio which might be an expression of the well-established cardiotoxicity of the drug.

Adenosine Triphosphate↗

PubMed1987

The effect of cytochrome c oxidase on lipid polymorphism of model membranes containing cardiolipin.

The effect of cytochrome c oxidase incorporation on the lipid polymorphism of the cardiolipin-Ca2+ system was investigated by 31P NMR and freeze-fracture electron microscopy. The integral membrane protein has a stabilizing effect on the bilayer organization of cardiolipin, in that it inhibits the Ca2+-induced HII phase formation of this lipid for Ca2+/cardiolipin molar ratios of 1-10. At a Ca2+/cardiolipin molar ratio of 25, about 80% of the lipid is organized in the HII phase and a structural phase separation occurs between the cardiolipin-Ca2+ complex organized in the hexagonal HII phase without protein and bilayer structures with incorporated protein.

Animals↗

PubMed1987

2H-NMR, 31P-NMR and DSC characterization of a novel lipid organization in calcium-dioleoylphosphatidate membranes. Implications for the mechanism of the phosphatidate calcium transmembrane shuttle.

2H-NMR, 31P-NMR and DSC investigations are presented on the structure and dynamics of the Ca2+-dioleoylphosphatidate complex which is formed upon addition of calcium to dispersions of pure dioleoylphosphatidate or of dioleoylphosphatidate in mixtures with dioleoylphosphatidylcholine (DOPC). It is concluded that the phosphate region in the polar headgroup of dioleoylphosphatidate is immobilized, while the oleate chains remain liquid and have increased disorder. In mixtures of dioleoylphosphatidate and DOPC in the presence of calcium a dioleoylphosphatidate-rich phase is segregated, in which the molecular behaviour of phosphatidate is rather similar to that of the pure Ca2+-dioleoylphosphatidate complex. A hypothetical model is proposed for the structure of this complex and this is correlated with the dioleoylphosphatidate-mediated transmembrane transport of calcium (Smaal, E.B., Mandersloot, J.G., De Kruijff, B. and De Gier, J. (1986) Biochim. Biophys. Acta 860, 99-108). Data indicate that this transmembrane shuttle is an inverted organization of phosphatidate molecules enclosing calcium ions in an anhydrous core.

Biological Transport↗

PubMed1987

Phase separation and hexagonal HII phase formation by gramicidins A, B and C in dioleoylphosphatidylcholine model membranes. A study on the role of the tryptophan residues.

The role of the tryptophan-residues in gramicidin-induced HII phase formation was investigated in dioleoylphosphatidylcholine (DOPC) model membranes. 31P-NMR and small angle X-ray diffraction measurements showed, that gramicidin A and C (in which tryptophan-11 is replaced by tyrosine) induce a similar extent of HII phase formation, whereas for gramicidin B and synthetic analogs in which one tryptophan, either at position 9 or 11 is replaced by phenylalanine, a dramatic decrease of the HII phase inducing activity can be observed. Modification of all four tryptophans by means of formylation of the indole NH group leads to a complete block of HII phase formation. Sucrose density centrifugation experiments on the various peptide/lipid samples showed a quantitative incorporation of the peptide into the lipid. For all samples in a 1/10 molar ratio of peptide to lipid distinct bands were found, indicative of a phase separation. For the gramicidin A'/DOPC mixture these bands were analyzed and the macroscopic organization was determined by 31P-NMR and small-angle X-ray diffraction. The results demonstrate that a quantitative phase separation had occurred between a lamellar phase with a gramicidin/lipid ratio of 1/15 and a hexagonal HII phase, which is highly enriched in gramicidin. A study on the hydration properties of tryptophan-N-formylated gramicidin in mixtures with DOPC showed that this analog has a similar dehydrating effect on the lipid headgroup as the unmodified gramicidin. In addition both the hydration study and sucrose density centrifugation experiments showed that, like gramicidin also its analogs have a tendency to aggregate, but with differences in aggregation behaviour which seemed related to their HII phase inducing activity. It is proposed that the main driving force for HII phase formation is the tendency of gramicidin molecules to self-associate and organize into tubular structures such as found in the HII phase and that whether gramicidin (analogs) form these or other types of aggregates depends on their tertiary structure, which is determined by intra- as well as intermolecular aromatic-aromatic stacking interactions.

Gramicidin↗

PubMed1987

Preferential lipid association and mode of penetration of apocytochrome c in mixed model membranes as monitored by tryptophanyl fluorescence quenching using brominated phospholipids.

The fluorescence of the single tryptophan residue at position 59 in apocytochrome c, the biosynthetic precursor of the inner mitochondrial membrane protein cytochrome c, was studied in small unilamellar vesicles composed of phosphatidylserine (PS) and phosphatidylcholine (PC) with or without specifically Br-labelled acyl chains at the sn-2 position. The protein has a very high affinity for PS-containing vesicles (dissociation constant Kd less than 1 microM). From the relative quenching efficiency by the brominated phospholipids, it could be concluded that the protein specifically associates with the PS component in mixed vesicles and that maximal quenching occurred with phospholipids in which the bromine was present at the 6,7-position of the 2-acyl chain suggesting that (part of) the bound protein penetrates 7-8 A deep into the hydrophobic core of the bilayer.

Apoproteins↗

PubMed1987

Consequences of the interaction of calcium with dioleoylphosphatidate-containing model membranes: calcium-membrane and membrane-membrane interactions.

Calcium binds to dioleoylphosphatidate/dioleoylphosphatidylcholine (DOPA/DOPC) (20:80, mol%) multilamellar vesicles in the presence of a calcium ionophore with stoichiometry of about 0.6 nmol calcium per nmol phosphatidate and an apparent dissociation constant of about 1.7 mM. Experiments on the behaviour of monomolecular films at an air/water interface show that calcium-phosphatidate binding results in a decrease in the area of the polar region of the phosphatidate molecule, probably caused by headgroup dehydration and partial charge neutralization. At calcium concentration higher than about 3 mM calcium neutralizes the negatively charged membrane surface of DOPA/DOPC (20:80, mol%) large unilamellar vesicles, and vesicle aggregation is observed. At 10 mM of calcium this results in a low level of vesicle fusion. These observed processes are not attended with calcium-induced phosphatidylcholine transbilayer movement in the membranes of DOPA/DOPC (20:80, mol%) large unilamellar vesicles. When these findings are compared with the results of a previous study on the permeability behaviour of large unilamellar vesicles of the same phospholipid composition under comparable conditions (Smaal, E.B., Mandersloot, J.G., De Kruijff, B. and De Gier, J. (1986) Biochim. Biophys. Acta 860, 99-108) the following conclusions can be drawn. At low millimolar calcium concentrations (less than 2.5 mM) calcium does not occupy all the binding sites of the membrane, no membrane-membrane interactions are observed and a selective translocation of calcium and calcium-chelating anions is appearing. The mechanism of this translocation may be explained by the formation of uncharged dehydrated complexes of calcium, phosphatidate and calcium chelator, which can pass the membrane via transient occurring non-bilayer structures. Between 3 and 10 mM of calcium an a selective permeability increase of the vesicular membrane is found, which is not a consequence of vesicle fusion but apparently of vesicle aggregation, possibly causing packing defects in the membrane.

Calcimycin↗

PubMed1987

Calcium-induced changes in permeability of dioleoylphosphatidylcholine model membranes containing bovine heart cardiolipin.

At calcium concentrations up to about 4 mM a selective permeability increase of cardiolipin/dioleoylphosphatidylcholine (50:50, mol%) membranes for calcium and its chelator arsenazo III is observed. Under these conditions calcium does not occupy all the binding sites of cardiolipin at the membrane interface and no vesicle-vesicle interactions are found. Lowering of the cardiolipin content of the vesicles to 20 mol% extends the calcium concentration range in which a selective permeability for calcium and arsenazo III is appearing up to about 12 mM. We suggest that the observed selective permeability increase is caused by transient formation of inverted micellar structures in the membrane with cardiolipin as translocating membrane component for calcium and arsenazo III. At calcium concentrations of 4 mM and higher for 50 mol% cardiolipin-containing vesicles a general permeability increase is found together with calcium-cardiolipin binding in a 1:1 stoichiometry, vesicles aggregation and, above 8 mM of calcium, vesicle fusion. The loss of barrier function of the membrane under these conditions is correlated with vesicle aggregation and may be explained by a transition from a bilayer into a hexagonal HII organization of the phospholipids.

Animals↗

PubMed1987

2H-nuclear magnetic resonance investigations on phospholipid acyl chain order and dynamics in the gramicidin-induced hexagonal HII phase.

The following results are reported in this paper: The interaction of gramicidin with [11,11-2H2]dioleoylphosphatidylcholine (DOPC) and [11,11-2H2]dioleoylphosphatidylethanolamine (DOPE) at different stages of hydration was studied by 2H- and 31P-nuclear magnetic resonance. In the L alpha phase in excess water the acyl chains of phosphatidylethanolamine (PE) are more ordered than phosphatidylcholine (PC) most likely as the result of the lower headgroup hydration of the former lipid. In excess water gramicidin incorporation above 5 mol % in DOPC causes a bilayer----hexagonal HII phase change. In the HII phase acyl chain order is virtually unaffected by gramicidin but the peptide restricts the fast chain motions. At low water content gramicidin cannot induce the HII phase but it markedly decreases chain order in the DOPC bilayer. Increasing water content results in separation between a gramicidin-poor and a gramicidin-rich L alpha phase with decreased order of the entire lipid molecule. Further increase in hydration reverts at low gramicidin contents the phase separation and at high gramicidin contents results in a direct change of the disordered lamellar to the hexagonal HII phase. Gramicidin also promotes HII phase formation in the PE system but interacts much less strongly with PE than with PC. The results support our hypothesis that gramicidin, by a combination of strong intermolecular attraction forces and its pronounced cone shape, both involving the four tryptophans at the COOH-terminus, has a strong tendency to organize, with the appropriate lipid, in intramembranous cylindrical structures such as is found in the HII phase.

Deuterium↗

PubMed1986

Relation between lipid polymorphism and transbilayer movement of lipids in rat liver microsomes.

We have studied the effects of trinitrophenylation on the transbilayer movement of phosphatidylcholine and the macroscopic lipid structure in rat liver microsomal membranes. The transbilayer movement of phosphatidylcholine was investigated using the PC-specific transfer protein. 31P-NMR was employed to monitor the phospholipid organization in intact microsomal vesicles. The results indicate that modification of microsomes with trinitrobenzenesulfonic acid enhances the transbilayer movement of phosphatidylcholine at 4 degrees C. Furthermore, phosphatidylethanolamine headgroup trinitrophenylation in microsomes increases the isotropic component in the 31P-NMR spectra even at 4 degrees C, possibly representing the appearance of intermediate non-bilayer lipid structures. The observed parallel between these data suggests that phosphatidylethanolamine molecules in the microsomal membrane, probably in combination with a protein component, are able to destabilize the bilayer organization, thereby facilitating the transmembrane movement of phospholipids.

Animals↗

PubMed1986

Ethylene glycol causes acyl chain disordering in liquid-crystalline, unsaturated phospholipid model membranes, as measured by 2H NMR.

2H NMR has been used to probe the effects of ethylene glycol at the level of the acyl chains in liposomes prepared from dioleoylphosphatidic acid or dioleoylphosphatidylcholine, labeled with 2H at the 11-position of both oleic acid chains. Increasing concentrations of ethylene glycol lead to a proportional and substantial decrease in the quadrupolar splittings, measured from the 2H NMR spectra of both liposomal systems, indicative of acyl chain disordering.

Ethylene Glycol↗

PubMed1986

Dolichyl phosphate induces non-bilayer structures, vesicle fusion and transbilayer movement of lipids: a model membrane study.

The effect of dolichol and dolichyl phosphate on fusion between large unilamellar vesicles comprised of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) and 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) was studied using a fluorescence resonance energy transfer assay. The influence of dolichyl phosphate on the transbilayer movement of DOPC in multilamellar vesicles (MLV) and large unilamellar vesicles (LUV) composed of DOPC and DOPE (1:2) was investigated by using the phosphatidylcholine-specific transfer protein. 31P-NMR and freeze-fracture electron microscopy were employed to study the macroscopic organization of DOPC and DOPE containing model membranes in the absence or presence of dolichyl phosphate. The results indicate that both dolichol and dolichyl phosphate enhance vesicle fusion in a comparable and concentration-dependent way; the amount of exchangeable PC from MLVs is increased by dolichyl phosphate, probably as a result of fusion processes; dolichyl phosphate destabilizes the bilayer organization in MLVs comprised of DOPE and DOPC, resulting in the formation of hexagonal (HII) phase and 'lipidic' particles.

Diterpenes↗

PubMed1986

Consequences of the interaction of calcium with dioleoylphosphatidate-containing model membranes: changes in membrane permeability.

The permeability behaviour of dioleoylphosphatidate/dioleoylphosphatidylcholine (20:80, mol%) large unilamellar vesicles at low millimolar calcium concentrations is different for various solutes. Between 0.5 mM and 2.5 mM of calcium a selective influx of calcium and efflux of enclosed calcium chelating anions is observed. At higher calcium concentrations the membrane loses its barrier function for a large variety of solutes. These permeability increases are a specific consequence of calcium phosphatidate interactions, because control experiments in which calcium was replaced by magnesium or in which dioleoylphosphatidate was replaced by dioleoylphosphatidylglycerol showed under the same conditions no permeability changes. These results are discussed on the basis of various putative mechanistic models for phosphatidate-mediated calcium translocation across membranes. Furthermore a kinetical model is presented by which the observed selective calcium and calcium-chelator translocation can be explained.

Arsenazo III↗

PubMed1986

Cytofluorescence detection of adriamycin-mitochondria interactions in isolated, perfused rat heart.

The major side-effect of the anthracycline anti-tumor drug adriamycin is a specific, dose-dependent cardiotoxicity. Impairment of mitochondrial function has been suggested to play an important role in this toxicity. The present study addresses the question as to whether direct drug-mitochondria interactions occur in the isolated, perfused rat heart. To this aim, cytofluorescence microscopy experiments were performed on thin cryosections. To demonstrate the applicability of this technique it is shown that adriamycin bound to isolated rat liver and heart mitochondria can be visualized through its characteristic fluorescence. Longitudinal sections from heart tissue perfused with 50 microM adriamycin display two distinct cellular sites of drug accumulation, i.e., nuclei which exhibit very bright fluorescence and, in addition, mitochondria which become significantly labeled with the drug. The mitochondrial localization of adriamycin is confirmed independently by quantification of the drug content of the mitochondrial fraction after cell fractionation. These results are discussed in the light of the potential role of adriamycin-nuclei versus adriamycin-mitochondria interactions in the deterioration of heart performance.

Animals↗

PubMed1986

Preferential association of apocytochrome c with negatively charged phospholipids in mixed model membranes.

The mitochondrial precursor protein, apocytochrome c, binds to model membranes containing negatively charged phospholipids (Rietveld, A., Sijens, R., Verkleij, A.J. and Kruijff, B. (1983) EMBO J. 2, 907-913). In the present paper the effect of apocytochrome c on the lipid distribution in model membranes, consisting of neutral and acidic phospholipids, is examined. Both ESR and fluorescence energy transfer experiments show that the protein preferentially interacts with the negatively charged phospholipid in the mixed model membranes. Semi-quantitative analysis of the fluorescence energy transfer from the single tryptophan in apocytochrome c to the parinaric acid in phosphatidylserine or phosphatidylcholine in mixed bovine brain phosphatidylserine/egg phosphatidylcholine vesicles reveals and average donor-acceptor distance of 22-26 A and 26-30 A for phosphatidylserine and phosphatidylcholine, respectively. In addition, these experiments demonstrate that this preferential interaction does not induce the separation of large domains enriched in complexes of apocytochrome c with negatively charged phospholipids and domains enriched in neutral lipids.

Apoproteins↗

PubMed1986

Polyphosphoinositides undergo charge neutralization in the physiological pH range: a 31P-NMR study.

The charge state of phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate was determined as a function of pH by way of 31P-NMR spectroscopy. The pK values for the first protonation of the phosphomonoester residues in phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate were found to be 6.2 and 6.6, respectively, for the 4-phosphate moiety, and 7.7 for the 5-phosphate moiety.

Animals↗

PubMed1986

Apocytochrome c binding to negatively charged lipid dispersions studied by spin-label electron spin resonance.

The interaction of apocytochrome c with aqueous dispersions of phosphatidylserine from bovine spinal cord and with other negatively charged phospholipids has been studied as a function of pH and salt concentration by using spin-label electron spin resonance (ESR) spectroscopy and chemical binding assays. The ESR spectra of phospholipids spin-labeled at different positions on the sn-2 chain indicate a generalized decrease in mobility of the lipids, while the characteristic flexibility gradient toward the terminal methyl end of the chain is maintained, on binding of apocytochrome c to phosphatidylserine dispersions. This perturbation of the bulk lipid mobility or ordering is considerably greater than that observed on binding of cytochrome c. In addition, a second, more motionally restricted, lipid component is observed with lipids labeled close to the terminal methyl ends of the chains. This second component is not observed on binding of cytochrome c and can be taken as direct evidence for penetration of apocytochrome c into the lipid bilayer. It is less strongly motionally restricted than similar spectral components observed with integral membrane proteins and displays a steep flexibility gradient. The proportion of this second component increases with increasing protein-to-lipid ratio, but the stoichiometry per protein bound decreases from 4.5 lipids per 12 000-dalton protein at low protein contents to 2 lipids per protein at saturating amounts of protein. Apocytochrome c binding to phosphatidylserine dispersions decreases with increasing salt concentration from a saturation value corresponding to approximately 5 lipids per protein in the absence of salt to practically zero at 0.4 M NaCl.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗