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C J Restall

Publications and source records attributed to C J Restall.

9 recordsLinked to original sources
PubMed1989

The effects of calcium, temperature and phospholamban phosphorylation on the dynamics of the calcium-stimulated ATPase of canine cardiac sarcoplasmic reticulum.

Highly purified sarcoplasmic reticulum (SR) has been prepared from dog hearts and has been incubated with the triplet probe erythrosinyl isothiocyanate to specifically label the Ca2+-stimulated ATPase (Ca2+-ATPase) of the SR. The rotational mobility of the Ca2+-ATPase has been studied in this erythrosin-labelled SR using time-resolved phosphorescence polarization. Qualitatively, the mobility of the cardiac Ca2+-ATPase resembles that of skeletal muscle SR Ca2+-ATPase. Addition of Ca2+ to SR affects the mobility of the Ca2+-ATPase in a way consistent with a segment of the ATPase altering its orientation relative to the plane of the membrane. Phosphorylation of phospholamban in cardiac SR by the purified catalytic subunit of cAMP-dependent protein kinase, which is known to increase the activity of the Ca2+-ATPase by deinhibition, also alters measured anisotropy. The changes observed are not compatible with dissociation of the Ca2+-ATPase from phospholamban after the latter is phosphorylated. The data are more consistent with phospholamban associating with the Ca2+-ATPase following phosphorylation, or more complex models in which only the hydrophilic domain of phospholamban binds with and dissociates from the Ca2+-ATPase.

Animals↗
PubMed1986

Derivative spectroscopy of tryptophan fluorescence used to study conformational transitions in the (Ca2+ + Mg2+)-adenosine triphosphatase of sarcoplasmic reticulum.

Second and fourth derivatives have been calculated from the fluorescence emission spectra of N-acetyl tryptophanamide in solvents of varying polarity. It is demonstrated that the otherwise featureless fluorescence emission spectrum can be resolved into a series of discrete bands by the use of the derivative technique. These bands appear to have their origins in the transitions of electrons from the first excited singlet state back to the various vibrational levels of the ground state. The shifting of the fluorescence emission maximum to shorter wavelengths upon decreasing the solvent polarity is shown to be due to changes in the relative contributions of each of the bands combined with smaller changes in the band positions. Derivative spectra have also been obtained from the intrinsic tryptophan fluorescence of the (Ca2+ + Mg2+)-adenosine triphosphatase of sarcoplasmic reticulum membranes. A similar pattern of bands is observed to that found in the model system and is consistent with the majority of the tryptophan residues being located in hydrophobic environments. Addition of calcium ions to the protein results in enhancement of the protein fluorescence accompanied by a small and hitherto unseen blue-shift of the spectrum. The mechanistic implications of this finding are discussed in relation to the calcium transport function of the protein.

Animals↗
PubMed1985

Second-derivative infrared spectroscopic studies of the secondary structures of bacteriorhodopsin and Ca2+-ATPase.

The resolution of minor amide components in the infrared spectra of membrane proteins has, in the past, been limited by the small differences in frequency compared to the large half-widths of the bands that are assigned to different secondary conformations. Here, second-derivative calculations are used to resolve the relatively weak bands that are associated with the beta-sheet conformation and the vibrations of some amino acid side chains in the infrared spectra of bacteriorhodopsin and Ca2+-activated adenosine-5'-triphosphatase (Ca2+-ATPase). The spectra presented indicate that bacteriorhodopsin in the purple membrane contains an appreciable amount of beta structure in addition to the predominant alpha II-helical structure. Both sarcoplasmic reticulum and purified Ca2+-ATPase in native lipids contain alpha-helical and random coil conformations together with a small amount of beta structure. In 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) Ca2+-ATPase adopts a secondary conformation similar to that in the sarcoplasmic reticulum, and this structure is unaffected by the phospholipid phase transition. A shift to a predominantly random coil conformation is associated with solubilization of both bacteriorhodopsin and Ca2+-ATPase in 20% Triton X-100. Second-derivative analysis of the carbonyl stretching vibrations of DMPC bilayers indicates that below the phase-transition temperature (Tm) both bacteriorhodopsin and Ca2+-ATPase perturb the interface region such that the sn-2 carbonyls adopt a conformation similar to the sn-1 carbonyls. Above Tm, these integral proteins have no effect on the static order of the interface region, and the conformational inequivalence of the sn-1 and sn-2 carbonyls is similar to that found in a pure lipid bilayer.

Animals↗
PubMed1985

Conformational changes in the (Ca2+ + Mg2+)-ATPase of sarcoplasmic reticulum detected using phosphorescence polarization.

The technique of time-averaged phosphorescence has been used to study the interaction of calcium ions and ATP with the (Ca2+ + Mg2+)-ATPase in sarcoplasmic reticulum vesicles. The presence of excess calcium ions was found to cause a 20% decrease in the phosphorescence emission anisotropy. This is interpreted as being due to a conformational change in the protein and is supported by data from time-resolved phosphorescence measurements which also show a lowering of the anisotropy. This change in the decay of the emission anisotropy is associated with only minor changes in the rotational relaxation time of the protein and is again suggestive of a conformational change in the protein. In some cases ATP was also observed to lower the time-averaged phosphorescence anisotropy possibly via an interaction with the low-affinity regulatory site of the protein.

Adenosine Triphosphate↗
PubMed1979

The modulation of membrane fluidity by hydrogenation processes. III. The hydrogenation of biomembranes of spinach chloroplasts and a study of the effect of this on photosynthetic electron transport.

A method is reported for the in situ modification of the lipids of isolated spinach chloroplast membranes. The technique is based on a direct hydrogenation of the lipid double bonds in the presence of the catalyst, chlorotris(triphenylphosphine)rhodium (I). The pattern of hydrogenation achieved suggests that the catalyst distributes amongst all of the membranes. The polyunsaturated lipids within the membranes are hydrogenated at a faster rate and at an earlier stage than are the monoenoic lipids. Whilst addition of the catalyst to the chloroplast causes an initial 10--20% decrease in Hill activity, saturation of up to 40% of the double bonds present can be accomplished without causing further significant alterations in photosynthetic electron transport processes or marked morphological changes of the chloroplast structure as observed in the electron microscope.

Chloroplasts↗