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Biomedical subjects

Paul Murphy

Publications and source records attributed to Paul Murphy.

At least 19 recordsLinked to original sources
PubMed2005

The X-ray structure of the type II secretion system complex formed by the N-terminal domain of EpsE and the cytoplasmic domain of EpsL of Vibrio cholerae.

Gram-negative bacteria use type II secretion systems for the transport of virulence factors and hydrolytic enzymes through the outer membrane. These sophisticated multi-protein complexes reach from the pore in the outer membrane via the pseudopilins in the periplasm and a multi-protein inner-membrane sub-complex, to an ATPase in the cytoplasm. The human pathogen Vibrio cholerae uses such a secretion machinery, called the Eps-system, for the export of its major virulence factor cholera toxin into the intestinal tract of the human host. Here, we describe the 2.4 A structure of the hetero-tetrameric complex of the N-terminal domain of the ATPase EpsE and the cytoplasmic domain of the inner membrane protein EpsL, which constitute the major cytoplasmic components of the Eps-system. A stable fragment of EpsE in complex with the cytoplasmic domain of EpsL was identified via limited proteolysis and facilitated the crystallization of the complex. This first structure of a complex between two different proteins of the type II secretion system reveals that the N-terminal domain of EpsE and the cytoplasmic domain of EpsL form a hetero-tetramer, in which EpsL is the central dimer and EpsE binds on the periphery. The dimer of EpsL in this complex is very similar to the dimer seen in the crystal structure of the native cytoplasmic domain of EpsL, suggesting a possible physiological relevance despite a relatively small 675 A2 buried solvent accessible surface. The N-terminal domain of EpsE, which forms a compact domain with an alpha+beta-fold, places its helix alpha2 in a mostly hydrophobic cleft between domains II and III of EpsL burying 1700 A2 solvent accessible surface. This extensive interface involves several residues whose hydrophobic or charged nature is well conserved and is therefore likely to be of general importance in type II secretion systems.

Amino Acid Sequence↗
PubMed2004

Surface-exposed hemophilic mutations across the factor VIII C2 domain have variable effects on stability and binding activities.

Factor VIII (fVIII) is a plasma glycoprotein that functions as an essential cofactor in blood coagulation. Its carboxyl-terminal "C2" domain is responsible for binding to both activated platelet surfaces and von Willebrand factor. We characterized the effect of 20 hemophilia-associated missense mutations across this domain (that all occur in patients in vivo) on its stability and its binding activities. At least six of these mutations were severely destabilizing, and another four caused moderate destabilization and corresponding reductions in both binding functions. One mutant (A2201P) displayed a significant reduction in its membrane binding activity but normal von Willebrand factor binding, while two others (P2300S and R2304H) caused the opposite effect. Several mutations (including L2210P, V2223M, M2238V, and R2304C) displayed near wild-type stabilities and binding activities and may instead affect mRNA splicing or alternative properties or functions of the protein. This study demonstrated that von Willebrand factor and membrane binding activities can be uncoupled and uniquely disrupted by different mutations and that either effect can lead to similar reductions in clotting activity. It also illustrated how a heterogeneous genetic disorder causes diverse molecular phenotypes that result in similar disease states.

Cell Membrane↗
PubMed2004

Conformational effects on glycoside reactivity: study of the high reactive conformer of glucose.

The effect of conformation on glycoside reactivity was investigated by studying the hydrolysis of a selection of 3,6-anhydroglucosides as models for glucose in the highly reactive (1)C(4) conformation. Methyl 3,6-anhydro-beta-D-glucopyranoside was found to hydrolyze 200-400 times faster than methyl glucosides in the (4)C(1) conformation, while methyl 3,6-anhydro-beta-D-galactopyranoside, which is in the B(1,4) conformation, was less reactive than methyl beta-D-galactopyranoside. Methyl (3,6-anhydro-beta-D-glucopyranosyl)-(1 --> 6)-alpha-D-glucopyranoside, methyl (3,6-anhydro-alpha-D-glucopyranosyl)-(1 --> 6)-alpha-D-glucopyranosyl-(1 --> 6)-alpha-D-glucopyranoside, and methyl (3,6-anhydro-beta-D-glucopyranosyl)-(1 --> 6)-alpha-D-glucopyranosyl-(1 --> 6)-alpha-D-glucopyranoside were prepared and found to react selectively at the anhydro residue. The finding that (1)C(4) conformers of glucosides are highly reactive species is in accordance with and supports previous results showing that axial OH groups are less electron withdrawing than equatorial OH groups.

Carbohydrate Conformation↗
PubMed2004

Attachment styles and psychological profiles of child sex offenders in Ireland.

When 29 child sex offenders, 30 violent offenders, 30 nonviolent offenders, and 30 community controls were compared, a secure adult attachment style was 4 times less common in the child sex offender group than in any of the other three groups. Ninety-three percent of sex offenders had an insecure adult attachment style. Compared with community controls, the child sex offender group reported significantly lower levels of maternal and paternal care and significantly higher levels of maternal and paternal overprotection during their childhood. Compared with all three comparison groups, the child sexual offenders reported significantly more emotional loneliness and a more external locus of control. With respect to anger management, the child sexual offenders' profile more closely approximated those of nonviolent offenders and community controls than that of violent offenders.

Adult↗
PubMed2003

Reduction of sleep-disordered breathing after physostigmine.

The cholinesterase inhibitor physostigmine (PHYS) was investigated in a double-blind, placebo-controlled, randomized, crossover trial of 10 male patients with moderate to severe obstructive sleep apnea. PHYS (0.12 microg/minute/kg, 7-hour infusion) reduced mean apnea/hypopnea index (AHI) by 13.6 (95% confidence interval [CI], 2.2-25.1) corresponding to 21.4% (95% CI, -5.5 to 47.9) and increased minimum SaO2 by 8.7% (95% CI, -0.3 to 17.7) corresponding to 23.2% (95% CI, 4.8-41.3). During the last third of the night, coinciding with predicted plasma concentration steady state, non-REM sleep AHI decreased by 19.2 (95% CI, 0.1-38.3) or 14.9% (95% CI, -43.6 to 77.7) and REM AHI by 33.8 (95% CI, 13.7-54.0) or 67.5% (95% CI, 49.7-85.3). Mean total sleep time was reduced by 74 minutes (95% CI, 33.9-114.9), but patients with the least pronounced sleep shortening had the largest reduction of AHI (r = 0.73, p < 0.02). The nocturnal decline in heart rate was reduced by PHYS. Moreover, resting (early-night placebo heart rate) was positively correlated with proportional reduction of REM apnea index (r = 0.69, p < 0.02). Body mass index was negatively correlated with reduction of REM AHI (r = 0.77, p < 0.02). This, predominantly REM-related, reduction of obstructive sleep apnea after PHYS may provide a new treatment option if the effects are maintained in long-term studies.

Adult↗
PubMed2003

Automated prediction of CASP-5 structures using the Robetta server.

Robetta is a fully automated protein structure prediction server that uses the Rosetta fragment-insertion method. It combines template-based and de novo structure prediction methods in an attempt to produce high quality models that cover every residue of a submitted sequence. The first step in the procedure is the automatic detection of the locations of domains and selection of the appropriate modeling protocol for each domain. For domains matched to a homolog with an experimentally characterized structure by PSI-BLAST or Pcons2, Robetta uses a new alignment method, called K*Sync, to align the query sequence onto the parent structure. It then models the variable regions by allowing them to explore conformational space with fragments in fashion similar to the de novo protocol, but in the context of the template. When no structural homolog is available, domains are modeled with the Rosetta de novo protocol, which allows the full length of the domain to explore conformational space via fragment-insertion, producing a large decoy ensemble from which the final models are selected. The Robetta server produced quite reasonable predictions for targets in the recent CASP-5 and CAFASP-3 experiments, some of which were at the level of the best human predictions.

Algorithms↗