Seasonal changes of cadmium and copper levels in stem-boring larvae of Agapanthia villosoviridescens (Coleoptera) on salt marshes of the Westerschelde Estuary.
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Animals↗
Biomedical subjects
M A Hemminga
Publications and source records attributed to M A Hemminga.
Deuterium nuclear magnetic resonance investigation of bacteriophage M13 coat protein in dimyristoylphosphatidylcholine liposomes using palmitic acid as a probe.
The effect of incorporation of various amounts of M13 bacteriophage coat protein on the bilayer order and acyl chain motion in dimyristoylphosphatidylcholine (DMPC) liposomes has been investigated using deuterium NMR of specifically deuterated palmitic acid as a bilayer probe, phosphorus NMR and additional spin-label electron spin resonance (ESR). The secondary structure of the M13 coat protein in these bilayers was determined from circular dichroism spectra. Phosphorus NMR spectra of the mixed liposomes are characteristic for DMPC organized in bilayers, also after incorporation of various levels of M13 protein. Circular dichroism spectra of the coat protein indicate that the protein conformation is predominantly a beta-structure (more than 75%). Various incorporation levels of M13 coat protein do not affect the order of the deuterium-labelled positions along the acyl chain at the carbon-2, 9 and 16 positions. In contrast, the spin-spin relaxation times decrease at higher protein levels, especially at the carbon-16 position. The spin-label ESR spectra of the same system using 14-doxylstearic acid as a label show a second, motionally restricted component, that is not observed by deuterium NMR. The NMR and ESR results are consistent with a model in which the fatty acid molecules are in a fast two-site exchange (at a rate of approx. 10(7) Hz) between the sites in the bulk of the lipid bilayer and the motionally restricted sites on the coat protein.
Capsid↗
Spin-label electron spin resonance study of bacteriophage M13 coat protein incorporation into mixed lipid bilayers.
The major coat protein of bacteriophage M13 was incorporated in mixed dimyristoylphosphatidylcholine/dimyristoylphosphatidylglycerol (80/20 w/w) vesicles probed with different spin-labeled phospholipids, labeled on the C-14 atom of the sn-2 chain. The specificity for a series of phospholipids was determined from a motionally restricted component seen in the electron spin resonance (ESR) spectra of vesicles with the coat protein incorporated. At 30 degrees C and pH 8, the fraction of motionally restricted phosphatidic acid spin-label is 0.36, 0.52, and 0.72 for lipid/protein ratios of 18, 14, and 9 mol/mol, respectively. The ESR spectra, analyzed by digital subtraction, resulted in a phospholipid preference following the pattern cardiolipin = phosphatidic acid greater than stearic acid = phosphatidylserine = phosphatidylglycerol greater than phosphatidylcholine = phosphatidylethanolamine. The specificities found are related to the composition of the target Escherichia coli cytoplasmic membrane.
Coliphages↗
Time-resolved tryptophan fluorescence anisotropy investigation of bacteriophage M13 coat protein in micelles and mixed bilayers.
Coat protein of bacteriophage M13 is examined in micelles and vesicles by time-resolved tryptophan fluorescence and anisotropy decay measurements and circular dichroism experiments. Circular dichroism indicates that the coat protein has alpha-helix (60%) and beta-structure (28%) in 700 mM sodium dodecyl sulfate micelles and predominantly beta-structure (94%) in mixed dimyristoylphosphatidylcholine/dimyristoylphosphatidic acid (80/20 w/w) small unilamellar vesicles. The fluorescence decay at 344 nm of the single tryptophan in the coat protein after excitation at 295 or 300 nm is a triple exponential. In the micelles the anisotropy decay is a double exponential. A short, temperature-independent correlation time of 0.5 +/- 0.2 ns reflects a rapid depolarization process within the coat protein. The overall rotation of the coat protein-detergent complex is observed in the decay as a longer correlation time of 9.8 +/- 0.5 ns (at 20 degrees C) and has a temperature dependence that satisfies the Stokes-Einstein relation. In vesicles at all lipid to protein molar ratios in the range from 20 to 410, the calculated order parameter is constant with a value of 0.7 +/- 0.1 from 10 to 40 degrees C, although the lipids undergo the gel to liquid-crystalline phase transition. The longer correlation time decreases gradually on increasing temperature. This effect probably arises from an increasing segmental mobility within the coat protein. The results are consistent with a model in which the coat protein has a beta-structure and the tryptophan indole rings do not experience the motion of the lipids in the bilayer because of protein-protein aggregation.
Circular Dichroism↗
Role of the N-terminal part of the coat protein in the assembly of cowpea chlorotic mottle virus. A 500 MHz proton nuclear magnetic resonance study and structural calculations.
The interaction of the oligonucleotides (Ap)8A and (A-T)5 with empty capsids of the coat protein of cowpea chlorotic mottle virus (CCMV) has been studied with 500 MHz 1H nuclear magnetic resonance. It is found that these oligonucleotides specifically bind to the arginine and lysine residues of the N-terminal arm of the protein. Upon this binding, immobilization of part of the N-terminal arm occurs. In addition, secondary structure predictions and energy calculations have been performed on the N-terminal arm. These calculations were carried out as a function of the charges on the arginine and lysine side-chains. For free coat protein, where the arginine and lysine side-chains are charged, the arm is found in a random-coil conformation. In the neutralized state, as for the coat protein in the virus, the arm adopts an alpha-helical conformation. The results support a previously published model for the assembly of CCMV, in which a random-coil to alpha-helix conformational transition, induced by neutralizing the arginine and lysine side-chains, plays an essential role.
Amino Acid Sequence↗
Calcium-stimulated myofibrillar ATPase activity correlates with shortening velocity of muscle fibres in Xenopus laevis.
The iliofibularis muscle of Xenopus laevis is reported to contain five types of fibres which have different force-velocity relationships. Ten fibres of each type were selected on the basis of succinate dehydrogenase activity, cross-sectional area and location in the muscle, in order to assess the validity of the fibre type classification. Maximum calcium-stimulated myofibrillar ATPase activity (Vmax) and apparent Michaelis constant (Km) for ATP were determined for these 50 fibres from serial sections. The values obtained varied according to the type of fibre. Type 1 had the highest and type 5 the lowest values for Km and Vmax. In a separate experiment, single freeze-dried fibres were used to determine the relationship between their ATP content and apparent Km for ATP. There was a tendency for high ATP concentrations in fibres with high Km values. When myofibrillar ATPase activity was related to the maximum velocity of shortening of the five fibre types, a significant correlation was found. It is concluded that calcium-stimulated myofibrillar ATPase histochemistry allows an estimate of the maximum shortening velocity of muscle fibres from Xenopus laevis.
Adenosine Triphosphatases↗
Direct effects of the hyperglycemic factor of the freshwater snail Lymnaea stagnalis on isolated glycogen cells.
In the freshwater pulmonate snail Lymnaea stagnalis special cells, the glycogen cells (GC) are present for the storage of glycogen reserves. These cells occur in large numbers in the anterior mantle region. In a previous paper in vitro experiments with intact anterior mantle tissue indicated that a hyperglycemic factor is released from the central nervous system (CNS) which stimulates glycogen mobilization in mantle tissue (M. A. Hemminga, J. J. Maaskant, W. Koomen, and J. Joosse (1985). Neuroendocrine control of glycogen mobilization in the freshwater snail Lymnaea stagnalis. Gen. Comp. Endocrinol. 57, 117-123). In the present study the question of whether this factor affects glycogen metabolism of GC isolated from mantle tissue was investigated. It is reported that in high-K+ Ringers the CNS is stimulated to release a factor which, in a dose-dependent way, inhibits glycogen synthesis in isolated GC (measured as a decreased incorporation of [14C]glucose into glycogen). Simultaneous with this glycogen synthesis-inhibiting effect, stimulation of glycogen degradation is found (measured as a decreased retention of prelabeled glycogen). It is concluded that (1) the factor released by the CNS having these effects on isolated GC is the same as the hyperglycemic factor which was reported to stimulate glycogen mobilization in intact mantle tissue, and (2) GC after isolation from mantle tissue have retained their ability to respond to this factor.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Neuroendocrine control of glycogen mobilization in the freshwater snail Lymnaea stagnalis.
In the freshwater snail Lymnaea stagnalis the anterior mantle region, which mainly consists of large numbers of special glycogen-storing cells, is an important depot for the energy reserves of this snail. In organ culture experiments the central nervous system (CNS), in contrast to other tissues, both inhibits glycogen synthesis (measured as incorporation of [14C]glucose into glycogen) and stimulates glycogen breakdown in anterior mantle tissue (measured as a decreased retention of prelabeled glycogen). These effects are dose dependent, with saturation at the highest doses tested. High-potassium Ringer solution stimulates the secretion of a CNS factor which induces glucose release by anterior mantle tissue. This glucose-release-stimulating effect of the CNS is also dose dependent, but saturation of the response was not achieved. It is concluded that inhibition of glycogen synthesis and stimulation of glycogen breakdown and glucose release are probably effects of a single neurohormone which controls glycogen mobilization from the storage cells in the mantle. Like similar factors in other animal phyla, this putative neurohormone is referred to as a hyperglycemic factor.
Animals↗
The hyperglycemic factor of the CNS of the freshwater snail Lymnaea stagnalis: interaction with glucose stimulation of glycogen synthesis and evidence for its release during anaerobiosis.
In the freshwater pulmonate snail Lymnaea stagnalis glycogen synthesis in the glycogen storage cells (GC) is stimulated by increasing external glucose concentrations. On the other hand, a hyperglycemic factor from the CNS inhibits glycogen synthesis and stimulates glycogen breakdown in these cells. This CNS factor may be involved in the physiological control of glycogen mobilization. In the present study the interaction between the inhibiting effect of the CNS factor and the stimulating effect of increasing glucose concentrations on glycogen synthesis in isolated GC was investigated. The results, analyzed by determination of the Michaelis-Menten parameters for saturation kinetics, suggest that the CNS factor increases the glucose concentration required for half-maximal stimulation of glycogen synthesis, but does not influence the maximum rate of synthesis (competitive type of inhibition). The role of the hyperglycemic factor during conditions of glycogen breakdown was investigated by extirpation of the cerebral ganglia (-CG), which contain the main release sites of the factor. Extirpation did not affect the hemolymph glucose concentration or the glycogen levels in the GC during starvation. However, -CG animals showed reduced levels of the hyperglycemia normally associated with exposure to anaerobic conditions.
Anaerobiosis↗
Magic angle spinning carbon-13 NMR of tobacco mosaic virus. An application of the high-resolution solid-state NMR spectroscopy to very large biological systems.
Magic angle spinning 13C NMR was used to study tobacco mosaic virus (TMV) in solution. Well-resolved 13C NMR spectra were obtained, in which several carbon resonances of amino acids of the TMV coat protein subunits that are not observable by conventional high-resolution NMR spectroscopy can be designed. RNA resonance were absent, however, in the magic angle spinning 13C NMR spectra. Since three different binding sites are available for each nucleotide of the RNA, this is probably due to a line broadening caused by distributions of isotropic chemical shift values. In 13C-enriched TM 13C-13C dipolar interactions also gave rise to line broadening. By suitable pulse techniques that discriminate carbon resonances on the basis of their T1 and T1 rho values, it was possible to select particular groups of carbon nuclei with characteristic motional properties. Magic angle spinning 13C NMR spectra obtained with these pulse techniques are extremely well resolved.
Carbon Isotopes↗
Electron-paramagnetic-resonance spectroscopy of spin-labelled cowpea-chlorotic-mottle virus.
Cowpea chlorotic mottle virus has been labelled with a maleimide spin label when the nucleoprotein particles were in the swollen state. After the spin-labelling reaction, the virus was dialyzed to bring the nucleoprotein particles into different conformational states. The mobility of the labelled part of the protein was monitored by electron paramagnetic resonance (EPR) spectroscopy and by the saturation-transfer EPR technique. The latter technique especially showed sensitivity to conditions that influence the RNA conformation and the RNA-protein interaction. The shape of the EPR and saturation-transfer EPR spectra can only be explained by an anisotropic motion of the whole spin-labelled viral protein or the spin-labelled part of the protein subunit in the core of the virus.
Electron Spin Resonance Spectroscopy↗
Carbon-13 nuclear magnetic resonance studies on tobacco mosaic virus and its protein.
Fourier transform nuclear magnetic resonance studies on 12% 13C-enriched tobacco mosaic virus (TMV) and its rod-like protein oligomers in solution with molecular weights up to 42 X 10(6) are reported. In the virus approximately 17% of the carbons of the protein subunit have line widths of less than or equal to 300 Hz and T1 less than or equal to 1 s and are concluded to be mobile with more than one degree of freedom of internal rotation about a carbon--carbon bond. In the rodlike polymer of TMV protein at pH 5.3, 30% of the carbons are mobile, which implies rotational motions about carbon--carbon bonds and/or motions of the protein subunits within the polymer. The presence of internal mobility is supported by the observation that 20% of the carbons in the double disklike oligomer show decreasing line width upon increasing temperature; the remaining resonances have line widths which are temperature independent during the double disklike polymerization process. Since the molecular weight of TMV protein polymers increases with increasing temperature, this demonstrates that all nuclei within the double dislike oligomer are mobile. NMR and X-ray data on the double disklike polymer reveal differences with respect to internal mobility.
Fourier Analysis↗
A spin label EPR study of tobacco mosaic virus protein.
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Electron Spin Resonance Spectroscopy↗
Saturation transfer electron paramagnetic resonance spectroscopy of spin labeled tobacco mosaic virus protein.
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Electron Spin Resonance Spectroscopy↗
Lipid-protein interactions in model membranes from bovine brain white matter. An ESR spin label and electron microscopy study.
Lipid-protein model membranes, prepared from bovine brain white matter and containing all the lipids and Folch-Lees proteolipids, have been studied in macroscopically oriented multibilayers. To examine the lipid environment the membranes were spin labeled with the cholestane spin label (3'-spiro(2'=(N-oxyl-4',4'-dimethyl-oxazolidine))5alpha-cholestane) and a fatty acid spin label (4',4'-dimethyloxazolidine-N-oxyl derivative of 5-ketostearic acid). The ESR spectra exhibit two components arising from fairly well oriented and completely unoriented lipids. Up to a temperature of 55 degrees C the amount of oriented lipids is almost constant, being about 35%. At higher temperatures this percentage drops rapidly to zero. It is shown that the presence of unoriented lipids arises mainly from disrupted areas in the lipid bilayer structure. This is confirmed by electron miccroscopy and from an analysis of the temperature dependence of the order parameters of the spin labels. The presence of locally disrupted lipid parts in the bilayer is discussed in relation to the interaction of the brain white matter lipids with Folch-Lees protein.
Animals↗
Spin-labelled AMP - an activator of phosphorylase.
1. A spin-labelled AMP derivative and its diamagnetic analogue activate phosphorylase b in the same way, but do not activate phosphorylase a. 2. The electron-spin-resonance spectra of the spin-labelled AMP derivative bound to phosphorylase b and a have "powderlike" characteristics indicating that the spin label is immobilised on the protein. From changes in the electron-spin-resonance spectrum of spin-labelled AMP as phosphorylase b or a is added, the dissociation constants were calculated. 3. The interactions of spin-labelled AMP and the diamagnetic analogue with phosphorylase b and a have been monitored by observing changes in the spectral properties of fluorescent and spin-label probes covalently attached to the enzyme. 4. The dissociation constants of spin-labelled AMP and phosphorylase b or a are 175 +/- 25 muM and 15 +/- 5 muM respectively. Similar dissociation constants are obtained for the diamagnetic analogue. The effect of these AMP derivatives on the covalently attached probe groups and on phosphorylase activity is compared to the effect of AMP and IMP.
Adenosine Monophosphate↗
Application of 31P NMR to model and biological membrane systems.
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Animals↗
An ESR Study of the mobility of the cholestane spin label in oriented lecithin-cholesterol multibilayers.
The motion of the cholestane spin label in oriented lecithin-cholesterol multibilayers is described in terms of a rotational diffusion about the long molecular axis with diffusion coefficient D parrell and a restricted random librational motion about axes perpendicular to the long axis with diffusion coefficient D1. The diffusion coefficients have been determined from the angular dependence of the ESR line shape at various temperatures and cholesterol contents. The temperature dependence of D parrell and D1 clearly shows the transition from the gel to liquid crystalline phase. Increasing amounts of cholesterol reduce the transition temperature. A strong reduction is found from o to 10 mole % cholesterol. At 50 mole % no longer a sharp transition is observed. In the temperature range from 40 to 80 degrees C the range of D is about 10 times larger than the range of D parrell, indicating a high activation energy for the librational motion arising from a strong hindrance by interaction with surrounding molecules. Cholesterol contents up to 10-20 mole % give an increase of D parrell and D1, arising from strong decrease of the transition temperature in this range. Above 10-20 mole % a reduction of D parrell and D1 is found. However, the effect of cholesterol is much stronger on D1 than on D parrell. In the liquid crystalline phase at about 60 degrees C the effect of cholesterol on D parrell is even negligible, while D1 strongly changes. This indicates that in the liquid crystalline phase only the librational motion is influenced by cholesterol, due to a denser packing of the molecules in the bilayer.
Binding Sites↗