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P Bayley

Publications and source records attributed to P Bayley.

17 recordsLinked to original sources
PubMed2004

Over-expression of parvalbumin in transgenic mice rescues motoneurons from injury-induced cell death.

Following nerve injury in neonatal rats, a large proportion of motoneurons die, possibly as a consequence of an increase in vulnerability to the excitotoxic effects of glutamate. Calcium-dependent glutamate excitotoxicity is thought to play a significant role not only in injury-induced motoneuron death, but also in motoneuron degeneration in diseases such as amyotrophic lateral sclerosis (ALS). Motoneurons are particularly vulnerable to calcium influx following glutamate receptor activation, as they lack a number of calcium binding proteins, such as calbindin-D(28k) and parvalbumin. Therefore, it is possible that increasing the ability of motoneurons to buffer intracellular calcium may protect them from cell death and prevent the decline in motor function that usually occurs as a consequence of motoneuron loss. In this study we have tested this possibility by examining the effect of neonatal axotomy on motoneuron survival and muscle force production in normal and transgenic mice that over-express parvalbumin in their motoneurons.The sciatic nerve was crushed in one hindlimb of new-born transgenic and wildtype mice. The effect on motoneuron survival was assessed 8 weeks later by retrograde labelling of motoneurons innervating the tibialis anterior muscle. Following nerve injury in wildtype mice, only 20.2% (+/-2.2, S.E.M.; n=4) of injured motoneurons survive long term compared with 47.2% (+/-4.4, S.E.M.; n=4) in parvalbumin over-expressing mice. Surprisingly, this dramatic increase in motoneuron survival was not reflected in a significant improvement in muscle function, since 8 weeks after injury there was no improvement in either maximal twitch and tetanic force, or muscle weights.Thus, inducing spinal motoneurons to express parvalbumin protects a large proportion of motoneurons from injury-induced cell death, but this is not sufficient to restore muscle function.

Animals↗
PubMed2000

Leap-dynamics: efficient sampling of conformational space of proteins and peptides in solution.

A molecular simulation scheme, called Leap-dynamics, that provides efficient sampling of protein conformational space in solution is presented. The scheme is a combined approach using a fast sampling method, imposing conformational 'leaps' to force the system over energy barriers, and molecular dynamics (MD) for refinement. The presence of solvent is approximated by a potential of mean force depending on the solvent accessible surface area. The method has been successfully applied to N-acetyl-L-alanine-N-methylamide (alanine dipeptide), sampling experimentally observed conformations inaccessible to MD alone under the chosen conditions. The method predicts correctly the increased partial flexibility of the mutant Y35G compared to native bovine pancreatic trypsin inhibitor. In particular, the improvement over MD consists of the detection of conformational flexibility that corresponds closely to slow motions identified by nuclear magnetic resonance techniques.

Alanine↗
PubMed1999

Association of calmodulin with cytoskeletal structures at different stages of HeLa cell division, visualized by a calmodulin-EGFP fusion protein.

The fusion protein of calmodulin (CaM) with the enhanced green fluorescent protein EGFP has been expressed in a stably transfected HeLa cell line in order to visualise the localisation of calmodulin during the cell cycle on a continuous basis in live cells, and for immunofluorescence colocalisation with cytoskeletal structures. High-resolution images of CaM-EGFP in the mitotic apparatus show the characteristic strongly convoluted structure of the centrosome. CaM-EGFP also apparently associates with both polar and mitotic microtubules, and with a specific intracentrosomal structure. During cytokinesis, CaM-EGFP is also found decorating selected oriented filaments in close proximity to microtubules in the midbody region. In interphase cells, it is seen with filamentous and punctuate localisation at the nuclear envelope. The intensity and continuity of the CaM-EGFP images suggest that a significant fraction of the cellular calmodulin remains attached to cytoplasmic structures during the cell cycle.

Calmodulin↗
PubMed1994

Lack of effect of flumazenil and CGS 8216 on the anxiolytic-like properties of loreclezole.

Loreclezole is a novel antiepileptic that interacts in a unique way with the GABAA receptor complex. Its anticonvulsant effect in rats is reversed by benzodiazepine receptor partial inverse agonists, such as CGS 8216 but not by the competitive benzodiazepine receptor antagonist, flumazenil (Ro 15-1788). In this study loreclezole (30.0-50.0 mg/kg i.p.) was found to induce an anxiolytic-like effect in a rat conditioned emotional response test that was reversed neither by flumazenil (10.0 mg/kg) nor by CGS 8216 (2-phenylpyrazolo[4,3-c]quinolin-3(5H)-one, 0.3-3.0 mg/kg). The results suggest that the anxiolytic-like effects of loreclezole are not mediated by the benzodiazepine receptor.

Animals↗
PubMed1994

A comparison of the effects of the novel muscarinic receptor agonists L-689,660 and AF102B in tests of reference and working memory.

Four experiments compared the CNS effects of a novel M1/M3 receptor agonist L-689,660 with those of the M1/M3 muscarinic receptor agonist AF102B. In the mouse tail-flick test of antinociception (TF) the minimum effective doses to increase tail-flick latency (MED) of L-689,660 and AF102B were 0.03 mg/kg and 10.0 mg/kg, respectively. In a rat conditioned-suppression-of-drinking (CSD) test of reference memory, doses of 0.3 and 1.0 mg/kg L-689,660 and a dose of 5.0 mg/kg AF102B reversed a scopolamine-induced deficit in performance (0.6 mg/kg). Although there was a tendency for L-689,660 to reverse the scopolamine-induced (0.4 mg/kg) performance deficit in a rat delayed-matching-to-position (DMTP) test, the difference failed to reach statistical significance. In contrast, a 5.0 mg/kg dose of AF102B potentiated the scopolamine-induced deficit in choice accuracy and the number of trials completed on this task. In a response sensitivity (RS) test, chain-pulling rates were significantly decreased by L-689,660 (MED = 0.03 mg/kg) and by AF102B (MED = 5.0 mg/kg). These results suggest that L-689,660 and AF102B may ameliorate or reverse a scopolamine-induced deficit, but only at doses that also reduce chain-pulling rates on operant schedules of reinforcement.

Animals↗
PubMed1989

A lateral cap model of microtubule dynamic instability.

The co-existence and interconversion of growing and shrinking microtubules have been termed 'dynamic instability,' and have been directly observed to occur under a variety of conditions in vitro and in vivo. Previous modeling was based on the concept of an extensive, fluctuating cap of tubulin-GTP to stabilise growing microtubules. A quantitative kinetic model is now presented in which only the terminal layer of the multi-start helical microtubule lattice contains tubulin-GTP molecules, comprising a 'Lateral Cap.' In Monte Carlo numerical simulation, this model readily produces the decisive experimental evidence of microtubule dynamics, and predicts a co-operative mechanism for microtubule transitions. The model also suggests how differing kinetic properties at opposite ends are the result of the intrinsic polarity of the microtubule lattice, reflecting the polarity of the tubulin alpha/beta heterodimer.

Guanosine Triphosphate↗
PubMed1988

Fast disassembly of microtubules induced by Mg2+ or Ca2+.

The extent and rate of disassembly of microtubules induced by the addition of high concentrations of magnesium and calcium have been measured. At 25 degrees C, the rate constant for microtubule disassembly increases more than ten-fold on increasing [Mg2+] from 4.0 to 20 mM. The process is even more sensitive to [Ca2+], showing similar enhancement on increasing [Ca2+] from 0.5 to 8 mM. Electron microscopy indicates that the disassembly is an end-dependent process. Complete microtubule disassembly occurs at concentrations in excess of 10 and 2 mM for Mg2+ and Ca2+, respectively; this suggests the importance of binding to weak sites for both ions. The sensitivity to ionic composition explains the wide variations in the published values for k-, under varying conditions. The results indicate the potential range of microtubule disassembly rates which may be encountered under different conditions in vitro and in vivo. The highest values of k- (ca. 3000 s-1) would imply microtubule shortening rates in excess of 100 um per minute.

Animals↗
PubMed1988

The conformation of calmodulin: a substantial environmentally sensitive helical transition in Ca4-calmodulin with potential mechanistic function.

The conformation of Ca4-calmodulin in solution, as assessed by far-UV peptide circular dichroism, contains significantly less alpha-helix than the proposed X-ray crystal structure. We now show that Ca4-calmodulin adopts significant additional helical structure in solution in the presence of a helicogenic solvent (50%, v/v, aqueous 2,2,2-trifluoroethanol or 50%, v/v, methylpentane-5,5-diol). We suggest that the long continuous helix (residues 66-92 of the crystal structure) is not necessarily a normal feature of the calmodulin structure in solution, and may be due in part to the conditions of crystallisation. This result is supported by time-resolved tyrosine fluorescence anisotropy studies indicating that Ca4-calmodulin in solution is an essentially compact globular structure which undergoes isotropic rotational motion. We conclude that, under appropriate ionic and apolar environmental conditions, Ca4-calmodulin undergoes a substantial helical transition, which may involve residues in the central region of the molecule. Such a transition could have an important function in determining specificity and affinity in interactions of calmodulin with different target sequences of Ca2+-dependent regulatory enzymes.

Animals↗
PubMed1988

Effect of amino acid substitutions and deletions on the thermal stability, the pH stability and unfolding by urea of bovine calbindin D9k.

The influence of amino acid substitutions and deletions on the stability of bovine calbindin D9k, the smallest protein known with a pair of EF-hand calcium-binding sites, has been studied using circular dichroism and ultraviolet absorption spectroscopy. The five modifications are confined to one of the two Ca2+ -binding sites. The Ca2+-loaded forms of the wild-type and mutant calbindins are too stable to be significantly denatured by heating at 90 degrees C or by adding 8 M urea. For the Ca2+-free (apo) forms thermal unfolding appears to be only half complete at 90 degrees C, while denaturation is complete in 7-8 M urea. Four of the mutant proteins show reduced resistance towards unfolding by urea, but one of the modified proteins (Glu-17----Gln) shows an increased stability, presumably because of a reduced electrostatic repulsion in the native state. According to X-ray crystallographic data the OH group of the single tyrosine of calbindin (Tyr-13) is hydrogen-bonded to the carboxyl group of Glu-35, thus linking the two alpha helices flanking the N-terminal Ca2+ site. The pK of ionization of the Tyr-13 hydroxyl group was over 13 for calcium forms of the wild-type protein, between 12.3 and 12.8 for the calcium form of three mutants and between 11.5 and 11.7 for the apoproteins. Significant differences in pH stability between wild type and mutants were observed in the calcium forms, but were not apparent in the apo forms.

Amino Acids↗
PubMed1984

The kinetics of calcium binding to calmodulin: Quin 2 and ANS stopped-flow fluorescence studies.

The rate of calcium dissociation from bovine testis calmodulin was measured by fluorescence stopped-flow using the calcium indicator Quin 2 or the fluorescence probe 8-anilinonaphthalene sulphonate. Two processes are resolved with Quin 2 corresponding to dissociation from the high affinity sites (kdiss 2 to 9 s-1 for T = 11 to 28 degrees C) and from the low affinity sites (kdiss 293 to 550 s-1 for T = 11 to 19 degrees C). These rates and the activation parameters as determined for the slow process delta H not equal to = 59 +/- 10 kJ X mol-1 and delta S not equal to = 30 +/- 30 JK-1 X mol-1 are in good agreement with values determined from the 43Ca NMR exchange rates. These experiments provide confirmation that the calcium induced conformational change cannot be resolved kinetically from the calcium binding or dissociation, and by inference this conformational change is not a rate-limiting process in the function of calmodulin.

Aminoquinolines↗