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J Cavanagh

Publications and source records attributed to J Cavanagh.

At least 19 recordsLinked to original sources
PubMed2006

International spinal research trust research strategy. III: A discussion document.

STUDY DESIGN: Discussion document. OBJECTIVES/METHODS: To review the Research Strategy of the International Spinal Research Trust (ISRT), which identifies key areas of basic and clinical research that are likely to be beneficial in developing potential treatments for spinal cord injury for funding. This strategy is intended to both guide the programme of research towards areas of priority and stimulate discussion of the different avenues of research. This latest document has been developed to take into account the scientific progress in the 6 years since publication of the previous Research Strategy. RESULTS/DISCUSSION: The latest scientific developments in research designed to repair the spinal cord and restore function following injury and how they might impact on spinal cord injury research are highlighted.

Biomedical Research↗
PubMed2004

Relapse into mania or depression following lithium discontinuation: a 7-year follow-up.

OBJECTIVE: We aimed to assess whether outcome followed a worse course after acute lithium withdrawal. METHOD: Data were acquired by review of the clinical records on 14 cases and 28 controls matched for age, gender and time on lithium for about 7 years following discontinuation. RESULTS: Significant differences were found between cases and controls. Cases experienced more episodes of depression and total affective relapse. The log-rank test revealed a significantly lower survival probability in cases than controls (P < 0.0009). However, the increased risk of recurrence was largely confined to the interval immediately after lithium discontinuation. CONCLUSION: These results confirm that acute discontinuation of lithium leads to a high immediate relapse rate. Most of the excess morbidity over 7 years appears to be attributable to the first episode following discontinuation. Outcome was not worsened by discontinuation.

Adult↗
PubMed2001

The DNA-binding domain in the Bacillus subtilis transition-state regulator AbrB employs significant motion for promiscuous DNA recognition.

AbrB is a Bacillus subtilis protein responsible for regulating a diverse array of unrelated genes during periods of sub-optimal growth conditions. DNA binding by AbrB is unique in that sequence recognition is specific, yet no obvious consensus sequence of bound promoter regions is apparent. The N-terminal domain is a recently characterized representative of a novel class of DNA-binding proteins that possess a looped-hinge helix DNA-binding topology. Although the structural characterization of this DNA-binding topology contributed to an understanding of the architectural basis for recognition of DNA target sequences, specific mechanisms responsible for promiscuity in DNA sequence recognition still were not apparent. Analysis of (15)N backbone relaxation parameters shows that dynamic motion of regions directly linked to DNA binding show concerted motion on the microsecond-millisecond timescale. Furthermore, dynamic motion of the hinge region suggests that the DNA-binding region is capable of conformational orientations that allow it to accommodate DNA sequence variability in the cognate binding sites.

Amino Acid Sequence↗
PubMed2001

Autologous biological response modification of the gonadotropin receptor.

It is generally held with respect to heterotrimeric guanine nucleotide binding protein-coupled receptors that binding of ligand stabilizes a conformation of receptor that activates adenylyl cyclase. It is not formally appreciated if, in the case of G-protein-coupled receptors with large extracellular domains (ECDs), ECDs directly participate in the activation process. The large ECD of the glycoprotein hormone receptors (GPHRs) is 350 amino acids in length, composed of seven leucine-rich repeat domains, and necessary and sufficient for high affinity binding of the glycoprotein hormones. Peptide challenge experiments to identify regions in the follicle-stimulating hormone (FSH) receptor (FSHR) ECD that could bind its cognate ligand identified only a single synthetic peptide corresponding to residues 221-252, which replicated a leucine-rich repeat domain of the FSHR ECD and which had intrinsic activity. This peptide inhibited human FSH binding to the human FSHR (hFSHR) and also inhibited human FSH-induced signal transduction in Y-1 cells expressing recombinant hFSHR. The hFSHR-(221-252) domain was not accessible to anti-peptide antibody probes, suggesting that this domain resides at an interface between the hFSHR ECD and transmembrane domains. CD spectroscopy of the peptide in dodecyl phosphocholine micelles showed an increase in the ordered structure of the peptide. CD and NMR spectroscopies of the peptide in trifluoroethanol confirmed that hFSHR-(221-252) has the propensity to form ordered secondary structure. Importantly and consistent with the foregoing results, dodecyl phosphocholine induced a significant increase in the ordered secondary structure of the purified hFSHR ECD as well. These data provide biophysical evidence of the influence of environment on GPHR ECD subdomain secondary structure and identify a specific activation domain that can autologously modify GPHR activity.

Amino Acid Sequence↗
PubMed2001

Clinical and pathological features of a Parkinsonian syndrome in a family with an Ala53Thr alpha-synuclein mutation.

We describe an Australian family of Greek origin with a parkinsonian syndrome and an Ala53Thr alpha-synuclein gene mutation. Five of 9 siblings were affected, the average age of onset was 45 years, and the initial symptoms were variable, including resting tremor, bradykinesia, and gait disturbance, as previously described in families with the same point mutation. Affected family members responded well to levodopa, developed progressive cognitive impairment, and had a disease duration of 5 to 16 years. Pathologic features typical of idiopathic Parkinson's disease were found at autopsy. However, there were several additional features not previously reported in families with this gene mutation. These features included severe central hypoventilation, orthostatic hypotension, prominent myoclonus, and urinary incontinence. An abundance of alpha-synuclein-immunoreactive Lewy neurites were found in the brainstem pigmented nuclei, hippocampus, and temporal neocortex. The Lewy neurites were associated with temporal lobe vacuolation. Subcortical basal ganglia cell loss and gliosis were seen. These additional clinical and pathological features suggest that the Ala53Thr alpha-synuclein mutation can produce a more widespread disorder than found in typical idiopathic Parkinson's disease.

Adult↗
PubMed2000

Two-dimensional relayed-rotating-frame overhauser spectroscopy (1)H NMR experiments for the selective identification of 1,2-glycosidic linkages in polysaccharides.

This communication describes the use of two-dimensional relayed (TOCSY)-ROESY experiments for the rapid and selective identification of alpha/beta1,2-glycosidic linkages in polysaccharides. The method assists in the identification of cross-peaks in crowded regions of ROESY spectra by moving them to less congested areas. In addition, the appearance of the spectra provides information relating the location of the glycosidic linkage within the sequence of the glycan under study. Selection of solely the 1,2- linkages is achieved by appropriately tuning the duration of the TOCSY mixing period. The method is demonstrated both theoretically and experimentally for a variety of test case polysaccharides.

Carbohydrate Sequence↗
PubMed2000

May the driving force be with you--whatever it is.

Changes in the atomic coordinate fluctuations contribute to the entropy of biomolecular processes such as complex formation. Characterizations of such changes in proteins reveal that the response may be dramatically different between the backbone and the side chains, and further resolve enthalpy-entropy compensation at the molecular level.

Calmodulin↗
PubMed2000

Novel DNA binding domain and genetic regulation model of Bacillus subtilis transition state regulator abrB.

We have determined the high resolution NMR solution structure of the novel DNA binding domain of the Bacillus subtilis transition state regulator AbrB. Comparisons of the AbrB DNA binding domain with DNA binding proteins of known structure show that it is a member of a completely novel class of DNA recognition folds that employs a dimeric topology for cellular function. This new DNA binding conformation is referred to as the looped-hinge helix fold. Sequence homology investigations show that this DNA binding topology is found in other disparately related microbes. Structural analysis of the AbrB DNA binding domain together with bioanalytical and mutagenic data of full length AbrB allows us to construct a general model that describes the genetic regulation properties of AbrB.

Bacillus subtilis↗
PubMed1999

Millisecond-timescale motions contribute to the function of the bacterial response regulator protein Spo0F.

Protein backbones and side chains display varying degrees of flexibility, which allows many slightly different but related conformational substates to occur. Such fluctuations are known to differ in both timescale and magnitude, from rotation of methyl groups (nanoseconds) to the flipping of buried tyrosine rings (seconds). Because many mechanisms for protein function require conformational change, it has been proposed that some of these ground-state fluctuations are related to protein function. But exactly which aspects of motion are functionally relevant remains to be determined. Only a few examples so far exist where function can be correlated to structural fluctuations with known magnitude and timescale. As part of an investigation of the mechanism of action of the Bacillus subtilis response regulator SpoOF, we have explored the relationship between the motional characteristics and protein-protein interactions. Here we use a set of nuclear magnetic resonance 15N relaxation measurements to determine the relative timescales of SpoOF backbone fluctuations on the picosecond-to-millisecond timescale. We show that regions having motion on the millisecond timescale correlate with residues and surfaces that are known to be critical for protein-protein interactions.

Bacillus subtilis↗
PubMed1999

Temperature dependence of intramolecular dynamics of the basic leucine zipper of GCN4: implications for the entropy of association with DNA.

The basic leucine zipper domain of the yeast transcription factor GCN4 consists of a C-terminal leucine zipper and an N-terminal basic DNA-binding region that achieves a stable structure only after association with DNA. Backbone dynamics of a peptide encompassing the basic and leucine zipper bZip domain (residues 226-281) are described using NMR spectroscopy. The 15N longitudinal relaxation rates, 15N transverse relaxation rates, and {1H}-15N nuclear Overhauser effects were measured for the backbone amide nitrogen atoms at 290 K, 300 K, and 310 K. The relaxation data were interpreted using reduced spectral density mapping to determine values of the spectral density function, J(omega), at the frequencies 0, omegaN, and 0.87omegaH to characterize overall and intramolecular motions on picosecond-nanosecond timescales. To account for the temperature dependence of overall rotational diffusion, the J(0) values were normalized using Stoke's Law. At 310 K, the 13Calpha and 13CO chemical shifts in conjunction with the spectral density values indicate that the leucine zipper sequence forms a highly ordered alpha-helix, while the basic region populates an ensemble of highly dynamic transient structures with substantial helical character. The normalized values of J(0) and the values of J(0.87omegaH) for residues in the leucine zipper dimerization domain are independent of temperature. In contrast, residues in the basic region exhibit pronounced increases in the normalized J(0) and decreases in J(0.87omegaH) as temperature is decreased. A strong correlation exists between the temperature dependence of 13CO chemical shifts and of J(0.87omegaH). These results suggest that, for the basic region, lowering the temperature increases the population of transient helical conformations, and concomitantly reduces the amplitude or timescale of conformational fluctuations on picosecond-nanosecond timescales. Changes in the conformational dynamics of the peptide backbone of the basic region that accompany DNA binding contribute to the overall thermodynamics of complex formation. The change in backbone conformational entropy derived from NMR spin-relaxation data agrees well with the result calculated from calorimetric measurements. Restriction of the conformational space accessible to the basic region may significantly reduce the entropic cost associated with formation of the basic region helices consequent to DNA binding.

Amino Acid Sequence↗
PubMed1999

Novel Schizosaccharomyces pombe N-linked GalMan9GlcNAc isomers: role of the Golgi GMA12 galactosyltransferase in core glycan galactosylation.

Schizosaccharomyces pombe synthesizes very large N-linked galactomannans, which are elongated from the Man9GlcNAc2 core that remains after the trimming of three Glc residues from the Glc3Man9GlcNAc2 originally transferred from dolichyl pyrophosphate to nascent proteins in the endoplasmic reticulum. Prior to elongation of the galactomannan outer chain, the Man9GlcNAc2 core is modified into a family of Hex10-15GlcNAc2 structures by the addition of both Gal and Man residues (Ziegler et al. (1994) J. Biol. Chem., 269, 12527-12535). To understand the pathway of Man9GlcNAc2 modification, the Hex10GlcNAc-sized pool was isolated by Bio-Gel P-4 gel filtration from the endo H-released N-glycans of S.pombe glycoproteins. This pool yielded four major fractions, a, b, c, and g, on preparative high pH, anion exchange chromatography, that represented 10, 29, 46, and 13% of the total Hex10GlcNAc present, respectively. Structures of the glycan isomers present in each fraction were determined by one- and two-dimensional 1H NMR spectroscopy techniques. Fraction a is principally (approximately 93%) a Man10GlcNAc with a new alpha1,2-linked Man cap on the upper-arm of Man9GlcNAc. Fraction b contained two isomers of GalMan9GlcNAc in which an alpha1,2-linked terminal Gal had been added either to the upper (b1, 30%) or middle-arm (b2, 70%) of Man9GlcNAc. The gma12 - alpha1,2-galactosyltransferase-negative S. pombe strain (Chappell et al. (1994) Mol. Biol. Cell., 5, 519-528) did not make fraction b implying that the gma12p galactosyltransferase is responsible for synthesis of both isomers b1 and b2. Isomer c is Man10GlcNAc in which a new branching alpha1, 6-linked Man had been added to the lower-arm alpha1,3-linked core residue as found earlier in Saccharomyces cerevisiae and Pichia pastoris. Fraction g had less than molar stoichiometry of both Gal and Glc. The major isomer (g1, 85%) is the Man9GlcNAc core with an alpha1,3-linked branching Gal on the penultimate 2-O-substituted Man of the lower arm. This residue is also found on a novel O-linked oligosaccharide recently described in S.pombe; Manalpha1,2(Galalpha1, 3)Manalpha1,2Mannitol (Gemmill and Trimble (1999) Glycobiology, 9, 507-515). The second isomer (g2, 15%) is the partially processed Glc2Man9GlcNAc intermediate. Defining these Hex10GlcNAc structures provides a starting point for understanding the enzymology of N-linked galactomannan core heterogeneity seen on S.pombe glycoproteins.

Carbohydrate Conformation↗
PubMed1999

The JIP group of mitogen-activated protein kinase scaffold proteins.

Activation of the c-Jun NH(2)-terminal kinase (JNK) group of mitogen-activated protein (MAP) kinases is mediated by a protein kinase cascade. This signaling mechanism may be coordinated by the interaction of components of the protein kinase cascade with scaffold proteins. The JNK-interacting protein (JIP) group of scaffold proteins selectively mediates signaling by the mixed-lineage kinase (MLK)-->MAP kinase kinase 7 (MKK7)-->JNK pathway. The scaffold proteins JIP1 and JIP2 interact to form oligomeric complexes that accumulate in peripheral cytoplasmic projections extended at the cell surface. The JIP proteins function by aggregating components of a MAP kinase module (including MLK, MKK7, and JNK) and facilitate signal transmission by the protein kinase cascade.

Adaptor Proteins, Signal Transducing↗
PubMed1999

The MKK7 gene encodes a group of c-Jun NH2-terminal kinase kinases.

The c-Jun NH2-terminal protein kinase (JNK) is a member of the mitogen-activated protein kinase (MAPK) group and is an essential component of a signaling cascade that is activated by exposure of cells to environmental stress. JNK activation is regulated by phosphorylation on both Thr and Tyr residues by a dual-specificity MAPK kinase (MAPKK). Two MAPKKs, MKK4 and MKK7, have been identified as JNK activators. Genetic studies demonstrate that MKK4 and MKK7 serve nonredundant functions as activators of JNK in vivo. We report here the molecular cloning of the gene that encodes MKK7 and demonstrate that six isoforms are created by alternative splicing to generate a group of protein kinases with three different NH2 termini (alpha, beta, and gamma isoforms) and two different COOH termini (1 and 2 isoforms). The MKK7alpha isoforms lack an NH2-terminal extension that is present in the other MKK7 isoforms. This NH2-terminal extension binds directly to the MKK7 substrate JNK. Comparison of the activities of the MKK7 isoforms demonstrates that the MKK7alpha isoforms exhibit lower activity, but a higher level of inducible fold activation, than the corresponding MKK7beta and MKK7gamma isoforms. Immunofluorescence analysis demonstrates that these MKK7 isoforms are detected in both cytoplasmic and nuclear compartments of cultured cells. The presence of MKK7 in the nucleus was not, however, required for JNK activation in vivo. These data establish that the MKK4 and MKK7 genes encode a group of protein kinases with different biochemical properties that mediate activation of JNK in response to extracellular stimuli.

Alternative Splicing↗
PubMed1998

Characterization of interactions between a two-component response regulator, Spo0F, and its phosphatase, RapB.

The phosphorelay signal transduction pathway controls sporulation initiation in Bacillus subtilis. Transfer of a phosphoryl group from multiple kinases (KinA and KinB) through a single domain response regulator homologue (Spo0F), a phosphotransferase (Spo0B), and ultimately to a transcriptional regulator, (Spo0A) activates sporulation. Counteracting this response are phosphatases (RapA and RapB), which can short-circuit this phosphorelay via dephosphorylation of Spo0F. In vitro assays of RapB activity on phosphorylated Spo0F alanine-scanning mutants have been used to identify Spo0F residues critical for interactions between these proteins. The Spo0F surface comprised of the beta1-alpha1 loop and N-terminal half of helix alpha1 has the largest number of residues in which an alanine substitution leads to resistance or decreased sensitivity to RapB phosphatase activity. Other mutations desensitizing Spo0F to RapB are also located near the site of phosphorylation on the beta3-alpha3 and beta4-alpha4 loops. This surface is similar to but not the same as the surface identified for KinA and Spo0B interactions with Spo0F. Divalent metal ions were shown to be required for RapB activity, and this activity was insensitive to vanadate, suggesting that Rap phosphatases catalyze acyl phosphate hydrolysis by inducing conformational changes in phosphorylated Spo0F, which results in increased autodephosphorylation. Arginine 16 of Spo0F is proposed to play a role in catalysis, and similarities between the mechanisms for RapB catalyzed Spo0F approximately P hydrolysis and GAP (GTPase activating protein)-assisted GTP hydrolysis of Ras are discussed.

Alanine↗
PubMed1998

A mammalian scaffold complex that selectively mediates MAP kinase activation.

The c-Jun NH2-terminal kinase (JNK) group of mitogen-activated protein (MAP) kinases is activated by the exposure of cells to multiple forms of stress. A putative scaffold protein was identified that interacts with multiple components of the JNK signaling pathway, including the mixed-lineage group of MAP kinase kinase kinases (MLK), the MAP kinase kinase MKK7, and the MAP kinase JNK. This scaffold protein selectively enhanced JNK activation by the MLK signaling pathway. These data establish that a mammalian scaffold protein can mediate activation of a MAP kinase signaling pathway.

Animals↗
PubMed1998

Identification of communication networks in Spo0F: a model for phosphorylation-induced conformational change and implications for activation of multiple domain bacterial response regulators.

Fundamental to understanding the mechanism by which phosphorylation activates bacterial signal transduction response regulator proteins is the identification of regions and residues that are responsible for the phosphorylation-induced conformational change. Here we review results from structural and protein dynamics investigations, and combine them with mutagenesis studies on the response regulator protein SpoOF to suggest a model in which a network of buried and surface residues link surface regions required for protein:protein interactions to the site of phosphorylation. The network described for SpoOF may provide pathways through which information is transmitted from the site of phosphorylation, propagating a conformational change many angstroms away. The general applicability of the communication network model for all bacterial response regulator proteins is discussed.

Bacterial Proteins↗
PubMed1998

CO_H(N)CACB experiments for assigning backbone resonances in 13C/15N-labeled proteins.

A triple resonance NMR experiment, denoted CO_H(N)CACB, correlates 1HN and 13CO spins with the 13C alpha and 13C beta spins of adjacent amino acids. The pulse sequence in an 'out-and-back' design that starts with 1HN magnetization and transfers coherence via the 15N spin simultaneously to the 13CO and 13C alpha spins, followed by transfer to the 13C beta spin. Two versions of the sequence are presented: one in which the 13CO spins are frequency labeled during an incremented t1 evolution period prior to transfer of magnetization from the 13C alpha to the 13C beta resonances, and one in which the 13CO spins are frequency labeled in a constant-time manner during the coherence transfer to and from the 13C beta resonances. Because 13CO and 15N chemical shifts are largely uncorrelated, the technique will be especially useful when degeneracy in the 1Hn-15N chemical shifts hinders resonance assignment. The CO_H(N)CACB experiment is demonstrated using uniformly 13C/15N-labeled ubiquitin.

Carbon Isotopes↗