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P J Stephens

Publications and source records attributed to P J Stephens.

At least 55 records · Page 3Linked to original sources
PubMed1991

Differential effects of alcohols on the spike threshold of an identified motor axon in a crab (Pachygrapsus crassipes).

Observations were made on the fast bender excitor (FBE) axon in autotomized crab limbs bathed in salines made up with different alcohols. It has been shown previously that the presence of ethanol at a certain level causes a single action potential to generate additional spikes in the peripheral axon branches. The present study examines the level of different alcohols required to induce peripheral spike generation. For primary alcohols, increasing the molecular weight decreased the level of alcohol required to produce peripheral spike generation. The threshold level of 2-butanol was greater than 1-butanol, but less than tertiary-butanol. These results are explained in terms of the partition coefficient, so that an alcohol with a higher partition coefficient enters the lipid bilayer more readily, thus a lower threshold level of that alcohol is required in the saline to generate additional spikes.

Action Potentials↗
PubMed1991

Site-directed mutagenesis of Azotobacter vinelandii ferredoxin I. Changes in [4Fe-4S] cluster reduction potential and reactivity.

We have used site-directed mutagenesis to obtain two variants of Azotobacter vinelandii ferredoxin I (AvFdI), whose x-ray structures are now available. In the C20A protein, a ligand to the [4Fe-4S] cluster was removed whereas in the C24A mutant a free cysteine next to that cluster was removed. Like native FdI, both mutants contain one [4Fe-4S] cluster and one [3Fe-4S] cluster. The structure of C24A is very similar to that of native FdI, while the structure of C20A is rearranged in the region of the [4Fe-4S] cluster to allow it to use the free Cys-24 as a replacement ligand. Here we compare the properties of the native, C20A, and C24A proteins. Although all three proteins are O2 stable in vitro, the C20A protein is much less stable toward proteolysis than the other two in vivo. Spectroscopic results show that all three proteins exhibit the same general redox behavior during O2-oxidation and dithionite reduction. Electrochemical data show that the [3Fe-4S] clusters in all three proteins have the same pH-dependent reduction potentials (-425 mV versus SHE, pH 7.8), whereas the [4Fe-4S] cluster potentials vary over a approximately 150 mV range from -600 mV (C24A) to -647 mV (native) to -746 mV (C20A). Despite this variation in potential both the C20A and C24A proteins appear to be functional in vivo. Native FdI reacts with three equivalents of Fe(CN)3-(6) to form a paramagnetic species previously proposed to be a cysteinyl-disulfide radical. Neither the C20A nor the C24A variant undergoes this reaction, strongly suggesting that it involves the free Cys-24.

Amino Acid Sequence↗
PubMed1991

Circular dichroism and magnetic circular dichroism of Azotobacter vinelandii ferredoxin I.

Room temperature circular dichroism (CD) and low temperature magnetic circular dichroism (MCD) spectra of air-oxidized and dithionite-reduced Azotobacter vinelandii ferredoxin I (FdI), a [( 4Fe-4S]2+/1+, [3Fe-4S]1+/0) protein, are reported. Unlike the CD of oxidized FdI, the CD of dithionite-reduced FdI exhibits significant pH dependence, consistent with protonation-deprotonation at or near the cluster reduced: the [3Fe-4S] cluster. The MCD of reduced FdI, which originates in the paramagnetic reduced [3Fe-4S]0 cluster, is also pH-dependent. Detailed studies of the field dependence and temperature dependence of the MCD of oxidized and reduced FdI, in the latter case at pH 6.0 and 8.3, are reported. The low-field temperature dependence of the MCD of oxidized FdI, which originates in the paramagnetic oxidized [3Fe-4S]1+ cluster, establishes the absence of a significant population of excited electronic states of this cluster up to 60 K. The low-field temperature dependence of the MCD of reduced FdI establishes that the ground-state manifold of the reduced [3Fe-4S]0 cluster possesses S greater than or equal to 2 at both pH 6.0 and 8.3. Analysis, assuming S = 2 and an axial zero-field splitting Hamiltonian, leads to D = -2.0 and -3.5 cm-1 at pH 6.0 and 8.3, respectively. The site of the (de)protonation affecting the spectroscopic properties of the [3Fe-4S] cluster remains unknown.

Azotobacter↗
PubMed1991

A case of autoerotic asphyxia associated with multiplex paraphilia.

During the past 20 years, the sensational aspects of autoerotic fatalities have captured the attention of medical examiners, psychiatrists, law enforcement agents, and the public, as well as the individuals themselves who engage in these dangerous practices. Reports of deaths related to sexual asphyxia have been presented numerous times at national and international meetings and have been the topic of discussion on television talk shows and in the press. Autoerotic fatalities and all the sexual curiosities related to these activities have prompted death scene investigators to publish case reports, and even textbooks, on the subject. The case presented herein is one of a multicomponent paraphilia in which self-asphyxiation (autoasphyxiophilia) led to a fatal autoerotic event.

Accidents↗
PubMed1991

Cases from the aerospace medicine residents' teaching file. Case #42. An aviator with concentration deficit, Lyme disease organic diagnostic evaluation, and a somatoform disorder.

An aviator with concentration deficit, Lyme disease organic diagnostic evaluation, and a somatoform disorder. The clinical presentation, evaluation and diagnosis of an aviator being evaluated for vague cognitive deficits of episodic and long duration with a history of rigid ideation concerning Lyme disease. The patient was diagnosed as having an atypical somatoform disorder. The aeromedical disposition of this patient is also presented.

Adult↗
PubMed1990

Site-directed mutagenesis of Azotobacter vinelandii ferredoxin I: [Fe-S] cluster-driven protein rearrangement.

Azotobacter vinelandii ferredoxin I is a small protein that contains one [4Fe-4S] cluster and one [3Fe-4S] cluster. Recently the x-ray crystal structure has been redetermined and the fdxA gene, which encodes the protein, has been cloned and sequenced. Here we report the site-directed mutation of Cys-20, which is a ligand of the [4Fe-4S] cluster in the native protein, to alanine and the characterization of the protein product by x-ray crystallographic and spectroscopic methods. The data show that the mutant protein again contains one [4Fe-4S] cluster and one [3Fe-4S] cluster. The new [4Fe-4S] cluster obtains its fourth ligand from Cys-24, a free cysteine in the native structure. The formation of this [4Fe-4S] cluster drives rearrangement of the protein structure.

Azotobacter↗
PubMed1989

Identification of a Ni- and Fe-containing cluster in Rhodospirillum rubrum carbon monoxide dehydrogenase.

Methyl viologen-oxidized carbon monoxide dehydrogenase (CODH) from Rhodospirillum rubrum exhibits complex EPR. Comparison to EPR of oxidized apo-CODH (CODH from which Ni is lacking) leads to the identification of signals whose intensity is correlated with the presence of Ni. 61Ni labeling observably broadens the sharpest feature of these signals, as does 57Fe. R. rubrum CODH thus contains a cluster containing both Ni and Fe. The EPR associated with this cluster is unlike any EPR previously attributed to Ni-containing prosthetic groups in other CODH enzymes or Ni-containing hydrogenases. The CO-analogue, CN-, perturbs the EPR signals that are attributed to the Ni-Fe species.

Aldehyde Oxidoreductases↗
PubMed1989

The axial ligands of heme in cytochromes: a near-infrared magnetic circular dichroism study of yeast cytochromes c, c1, and b and spinach cytochrome f.

Room temperature near-infrared magnetic circular dichroism and low-temperature electron paramagnetic resonance measurements have been used to characterize the ligands of the heme iron in mitochondrial cytochromes c, c1, and b and in cytochrome f of the photosynthetic electron transport chain. The MCD data show that methionine is the sixth ligand of the heme of oxidized yeast cytochrome c1; the identify of this residue is inferred to be the single conserved methionine identified from a partial alignment of the available cytochrome c1 amino acid sequences. A different residue, which is most likely lysine, is the sixth heme ligand in oxidized spinach cytochrome f. The data for oxidized yeast cytochrome b are consistent with bis-histidine coordination of both hemes although the possibility that one of the hemes is ligated by histidine and lysine cannot be rigorously excluded. The neutral and alkaline forms of oxidized yeast cytochrome c have spectroscopic properties very similar to those of the horse heart proteins, and thus, by analogy, the sixth ligands are methionine and lysine, respectively.

Amino Acid Sequence↗
PubMed1989

Substitution of histidine for arginine-101 of dinitrogenase reductase disrupts electron transfer to dinitrogenase.

Dinitrogenase reductase from Klebsiella pneumoniae strain UN1041 has a histidine residue substituted for arginine at position 101. The mutant dinitrogenase reductase was purified and characterized in order to determine the importance of arginine-101 in the interaction between dinitrogenase and dinitrogenase reductase during electron transfer. Purified dinitrogenase reductase from UN1041 is a dimer of 67 kDa, contains a functional 4Fe-4S cluster, undergoes a MgATP-dependent conformational change, and is competent for ATP hydrolysis uncoupled from substrate reduction in the presence of dinitrogenase. However, the mutant protein is unable to support the reduction of protons or acetylene by dinitrogenase. A 100-fold molar excess of Kp2 from UN1041 does not inhibit electron transfer from wild-type dinitrogenase reductase to dinitrogenase. It is concluded that the interaction of dinitrogenase reductase with dinitrogenase during reductant-independent ATP hydrolysis is different than the interaction between the two proteins during electron transfer; the substitution of histidine for arginine at position 101 disrupts only the latter interaction. The same conclusions are reached using wild-type dinitrogenase reductase which has been ADP-ribosylated at arginine-101.

Adenosine Triphosphate↗
PubMed1988

An axonal spike in an identified fast crab motor neuron has sodium and calcium components.

1. High intensity intracellular stimulation of the FBE axon produced a spike discharge. The spikes divided into two components, each with a different frequency. 2. Ion replacement and blocking specific ion channels revealed that the FBE spike has calcium and sodium components. 3. The pronounced depolarizing wave that follows the FBE spike is not produced by changes in calcium conductance.

Action Potentials↗
PubMed1988

Nickel-deficient carbon monoxide dehydrogenase from Rhodospirillum rubrum: in vivo and in vitro activation by exogenous nickel.

An inactive, Ni-deficient form of carbon monoxide (CO) dehydrogenase [carbon-monoxide:(acceptor) oxidoreductase; EC 1.2.99.2], designated apo-CO dehydrogenase, accumulated in Rhodospirillum rubrum when cells were grown in the absence of Ni and treated with CO. In vivo, both CO dehydrogenase activity and hydrogenase activity increased several hundred fold upon addition of 2 microM NiCl2. Apo-CO dehydrogenase was purified to homogeneity and differed from holo-CO dehydrogenase only in its activity and Ni content, containing less than 0.2 mol of Ni per mol of protein, and a specific activity of 35 mumol of CO per min per mg. Optimal in vitro activation of purified apo-CO dehydrogenase resulted in an enzyme with a specific activity of 2640 mumol of CO per min per mg. No additional enzymes or low molecular weight cofactors were required for activation. Apo-CO dehydrogenase was not activated by MgCl2, MnCl2, CuCl2, ZnCl2, CoCl2, or Na2MoO4. 63Ni was incorporated into apo-CO dehydrogenase during activation. The electron paramagnetic resonance (EPR) spectra of dithionite-reduced apo- and holo-enzyme were identical, indicating that, in the reduced state, the Fe-S centers observed by EPR are unchanged in the apo-enzyme.

Aldehyde Oxidoreductases↗
PubMed1987

Rectal impaction following enema with concrete mix.

This article describes an unusual rectal foreign body resulting from homosexual anal erotic activities. The patient had used an enema containing a concrete mix which became impacted and required surgical removal. The use, abuse, and complications of enemas are reviewed.

Adult↗
PubMed1985

Selective oxidative destruction of iron-sulfur clusters. Ferricyanide oxidation of Azotobacter vinelandii ferredoxin I.

The destructive oxidation of aerobically isolated 7Fe Azotobacter vinelandii ferredoxin I [(7Fe)FdI] by Fe(CN)3-6 is examined using low-temperature magnetic circular dichroism (MCD) and EPR. The results demonstrate that oxidation of the [3Fe-3S] cluster occurs only after essentially complete destruction of the [4Fe-4S] cluster. It is therefore feasible by controlled Fe(CN)3-6 oxidation to obtain a partially metallated form of FdI, (3Fe)FdI, containing only a [3Fe-3S] cluster. The MCD and EPR data demonstrate that the [3Fe-3S] cluster in (3Fe)FdI is essentially identical in structure to that in the native protein.

Azotobacter↗
PubMed1985

Neuromuscular relationships in the abdomen of the Californian shore crab Pachygrapsus crassipes.

There are two pairs of muscles in each abdominal segment of the crab; one pair of flexors and one pair of extensors. In the early larval stages the muscles have short sarcomeres--a property of fast fibers--and high thin to thick filament ratios--a property of slow fibers. In the adult the abdominal muscles are intermediate and slow, since they have fibers with intermediate and long sarcomeres, high thin to thick filament ratios, low myofibrillar ATPase activity, and high NADH diaphorase activity. The different fiber types are regionally distributed within the flexor muscle. Microelectrode recordings from single flexor muscle fibers in the adult showed that most fibers are supplied by three excitatory motor axons, although some are supplied by as many as five efferents. One axon supplies all of the flexor muscle fibers in its own hemisegment, and the evoked junctional potentials exhibit depression. This feature together with the innervation patterns of the fibers are similar to those reported for the deep flexor muscles of crayfish and lobsters. Therefore, in the adult crab, the abdominal flexor muscles have some features in common with the slow superficial flexors of crayfish and other features in common with the fast deep flexor muscles.

Adenosine Triphosphatases↗
PubMed1985

Morphological and physiological properties of the giant interneuron of the hermit crab (Pagurus pollicaris).

The physiological and morphological properties of the giant interneurons in the hermit crab Pagurus pollicaris are described. The cell bodies are located anteriorly in the supraesophageal ganglion, close to the mid-line. Each cell sends a neurite posteriorly and then laterally, so that they cross over in the center of the ganglion. Each axon then branches: one branch runs laterally while the other travels posteriorly and leaves the ganglion in the circumesophageal connective on the side contralateral to the cell body. The giant axons travel in the circumesophageal connectives and through the thoracic and abdominal ganglia without branching. Each giant axon makes synaptic contact with its ipsilateral giant abdominal flexor motor neuron and with a second flexor motor neuron that has its axon in the contralateral third root. In the supraesophageal ganglion there is a bidirectional synapse between the two giant interneurons. Intracellular recordings from the giant axons show that there is a delay of 0.5 to 0.75 ms that cannot be accounted for by spike propagation along the axons, and may be accounted for by a chemical synapse between the giant interneurons.

Action Potentials↗
PubMed1985

Temperature effects on a slow-crustacean neuromuscular system.

The membrane potential of the E2 axon and the bender muscle fibers increased with temperature. The input resistance of the axon, the spike amplitude and time course declined with temperature. Excitatory junctional potentials (ejps) exhibited maximum amplitudes and minimum facilitation at about the same temperature. Ejp time course and muscle membrane input resistance declined with temperature. Tension produced by the muscle also declined but then increased when additional spikes were generated in the periphery of the E2 axon.

Animals↗