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Biomedical subjects

M O'Donnell

Publications and source records attributed to M O'Donnell.

At least 145 records · Page 8Linked to original sources
PubMed1996

Replication of a P50 auditory gating deficit in Australian patients with schizophrenia.

Schizophrenic patients reportedly have a deficit in the control of sensitivity to auditory stimuli as shown by the P50 auditory evoked potential wave in a conditioning-testing paradigm that measures suppression of response to a repeated stimulus. Although this finding has been replicated by several US laboratories, one European group has not found differences between schizophrenic patients and normal control subjects. In the present study, investigators in the Schizophrenia Research Center at the Prince of Wales Hospital in Sydney, Australia, selected 22 normal control subjects, 11 acutely ill schizophrenic inpatients, and 11 clinically stable schizophrenic outpatients. Both schizophrenic groups were treated with similar doses of classical neuroleptic medications. Evoked potentials were recorded by an investigator from the US laboratory that initially reported the difference; five averages, each the response to 32 stimulus pairs, were recorded from each subject. The normal control subjects demonstrated significantly more suppression of the P50 response to the repeated stimuli than the schizophrenic groups, as previously reported. There were no significant changes in the suppression measure over the five trials. The suppression of the P50 wave by schizophrenic outpatients was somewhat greater than that by schizophrenic inpatients, but both schizophrenic groups had decreased suppression, compared with the normal subjects. The mean P50 suppression for five averages was successfully used in a logistic regression to classify subjects as normal or schizophrenic. This method was more accurate than attempts to classify subjects with only one average. The mean amplitude of the initial conditioning response did not differ between groups. Schizophrenic patients had slightly shorter mean latencies. There was no direct relationship of P50 suppression to measures of clinical psychopathology.

Adult↗
PubMed1996

Replisome assembly reveals the basis for asymmetric function in leading and lagging strand replication.

The E. coli replicase, DNA polymerase III holoenzyme, contains two polymerases for replication of duplex DNA. The DNA strands are antiparallel requiring different modes of replicating the two strands: one is continuous (leading) while the other is discontinuous (lagging). The two polymerases within holoenzyme are generally thought to have asymmetric functions for replication of these two strands. This report finds that the two polymerases have equal properties, both are capable of replicating the more difficult lagging strand. Asymmetric action is, however, imposed by the helicase that encircles the lagging strand. The helicase contact defines the leading polymerase constraining it to a subset of actions, while leaving the other to cycle on the lagging strand. The symmetric actions of the two polymerases free holoenzyme to assemble into the replisome in either orientation without concern for a correct match to one or the other strand.

Bacterial Proteins↗
PubMed1996

Dynamics of DNA-tracking by two sliding-clamp proteins.

Bacteriophage T4 gene 45 protein (gp45) and Escherichia coli beta are DNA-tracking sliding-clamp proteins that increase processivity by tethering their conjugate DNA polymerases to DNA. gp45 also activates T4 late transcription. DNA loading of gp45 and beta requires ATP or dATP hydrolysis; efficient loading at primer-template junctions is assisted by single-stranded DNA-binding proteins. The kinetics of gp45 loading and tracking have been examined by DNase I footprinting of linear DNA with one blunt end, one primer-template junction, and binding sites for proteins that block gp45 tracking. DNA loading of gp45 can also be interrupted by adding the non-hydrolyzable ATP analog ATP-gamma-S. At saturation, DNA is very closely packed with gp45 or beta. When gp45 loading is interrupted, or when a segment of the track is blocked off, the gp45 footprint dissipates within seconds, but the DNA-tracking state of beta is much more stable. The stability of the tracking state of gp45 is, however, increased by the macromolecular crowding agent polyethylene glycol. We suggest that labile gp45 catenation directly generates the coupling of late transcription to DNA replication during bacteriophage T4 multiplication.

Adenosine Triphosphate↗
PubMed1996

In vitro reconstitution of human replication factor C from its five subunits.

Replication factor C (RFC, also called Activator I) is part of the processive eukaryotic DNA polymerase holoenzymes. The processive elongation of DNA chains requires that DNA polymerases are tethered to template DNA at primer ends. In eukaryotes the ring-shaped homotrimeric protein, proliferating cell nuclear antigen (PCNA), ensures tight template-polymerase interaction by encircling the DNA strand. Proliferating cell nuclear antigen is loaded onto DNA through the action of RFC in an ATP-dependent reaction. Human RFC is a protein complex consisting of five distinct subunits that migrate through SDS/polyacrylamide gels as protein bands of 140, 40, 38, 37, and 36 kDa. All five genes encoding the RFC subunits have been cloned and sequenced. A functionally identical RFC complex has been isolated from Saccharomyces cerevisiae and the deduced amino acid sequences among the corresponding human and yeast subunits are homologous. Here we report the expression of the five cloned human genes using an in vitro coupled transcription/translation system and show that the gene products form a complex resembling native RFC that is active in supporting an RFC-dependent replication reaction. Studies on the interactions between the five subunits suggest a cooperative mechanism in the assembly of the RFC complex. A three-subunit core complex, consisting of p36, p37, and p40, was identified and evidence is presented that p38 is essential for the interaction between this core complex and the large p140 subunit.

DNA Replication↗
PubMed1996

Reconstitution of repair-gap UV mutagenesis with purified proteins from Escherichia coli: a role for DNA polymerases III and II.

Using a cell-free system for UV mutagenesis, we have previously demonstrated the existence of a mutagenic pathway associated with nucleotide-excision repair gaps. Here, we report that this pathway can be reconstituted by using six purified proteins: UvrA, UvrB, UvrC, DNA helicase II, DNA polymerase III core, and DNA ligase. This establishes the minimal requirements for repair-gap UV mutagenesis. DNA polymerase II could replace DNA polymerase III, although less effectively, whereas DNA polymerase I, the major repair polymerase, could not. DNA sequence analysis of mutations generated in the in vitro reaction revealed a spectrum typical of mutations targeted to UV lesions. These observations suggest that repair-gap UV mutagenesis is performed by DNA polymerase III, and to a lesser extent by DNA polymerase II, by filling-in of a rare class of excision gaps that contain UV lesions.

Adenosine Triphosphatases↗
PubMed1996

A molecular switch in a replication machine defined by an internal competition for protein rings.

Replication machines use ring-shaped clamps that encircle DNA to tether the polymerase to the chromosome. The clamp is assembled on DNA by a clamp loader. This report shows that the polymerase and clamp loader coordinate their actions with the clamp by competing for it through overlapping binding sites. The competition is modulated by DNA. In the absence of DNA, the clamp associates with the clamp loader. But after the clamp is placed on DNA, the polymerase develops a tight grip on the clamp and out-competes the clamp loader. After replication of the template, the polymerase looses affinity for the clamp. Now the clamp loader regains access to the clamp and removes it from DNA thus recycling it for future use.

Binding, Competitive↗
PubMed1996

Absorbable skin graft staples: a clinical trial using Graftac-X.

Skin stapling devices are a quick, effective method for securing skin grafts in burns patients. Removal of staples can be painful, often requiring a general anaesthetic. This problem has led to the recent development of absorbable staples which extrude with time. A prospective controlled clinical trial was performed on 20 burns patients comparing absorbable skin tacks with stainless steel staples. An area of at least 50 cm2 was grafted in each patient, half of which was secured with absorbable tacks and half with stainless steel staples. Each patient therefore acted as his/her own control. The presence of infection, haematoma and graft take using a grided cellophane sheet were assessed at one week. Infection and excess scarring at tack sites were assessed at one month. A statistically significant difference was not found between either group. In our hands absorbable skin staples have proven to be a reliable method of securing skin grafts.

Adult↗
PubMed1996

Clamp loading, unloading and intrinsic stability of the PCNA, beta and gp45 sliding clamps of human, E. coli and T4 replicases.

BACKGROUND: The high speed and processivity of replicative DNA polymerases reside in a processivity factor which has been shown to be a ring-shaped protein. This protein ("sliding clamp') encircles DNA and tethers the catalytic unit to the template. Although in eukaryotic, prokaryotic and bacteriophage-T4 systems, the processivity factors are ring-shaped, they assume different oligomeric states. The Escherichia coli clamp (the beta subunit) is active as a dimer while the eukaryotic and T4 phage clamps (PCNA and gp45, respectively) are active as trimers. The clamp can not assemble itself on DNA. Instead, a protein complex known as a clamp loader utilizes ATP to assemble the ring around the primer-template. This study compares properties of the human PCNA clamp with those of E. coli and T4 phage. RESULTS: The PCNA ring is a stable trimer down to a concentration below 100 nM (Kd approximately 21 nM). On DNA, the PCNA clamp slides freely and dissociates from DNA slowly (t1/2 approximately 24 min). beta is more stable in solution (Kd < 60 PM) and on DNA (t1/2 approximately 1 h) than PCNA which may be explained by its simpler oligomeric state. The T4 gp45 clamp is a much less stable trimer than PCNA (Kd approximately 250 nM) and requires association with the polymerase to stabilize it on DNA as observed previously. The consequence of this cooperation between clamp and polymerase is that upon finishing a template and dissociation of the polymerase from DNA, the gp45 clamp spontaneously dissociates from DNA without assistance. However, the greater stability of the PCNA and beta clamps on DNA necessitates an active process for their removal. The clamp loaders (RFC and gamma complex) were also capable of unloading their respective clamps from DNA in the presence of ATP. CONCLUSIONS: The stability of the different clamps in solution correlates with their stability on DNA. Thus, the low stability of the T4 clamp explains the inability to isolate gp45 on DNA. The stability of the PCNA and beta clamps predicts they will require an unloading factor to recycle them on and off DNA during replication. The clamp loaders of PCNA and beta double as clamp unloaders presumably for the purpose of clamp recycling.

Bacteriophage T4↗
PubMed1996

Neural tube defects 1974-94--down but not out.

AIMS: To describe accurately the total prevalence of neural tube defects (NTDs) in England and Wales over time, and to provide a benchmark up to 1994. METHODS: National data about NTDs reported as births or terminations are available from 1974-94, but reporting is incomplete. A local register of NTDs covering Oxfordshire/west Berkshire from 1965-94 was used to validate national data for the locality, using the method of capture and recapture, and hence to estimate incompleteness of reporting nationally. RESULTS: National underreporting is consistent at about two thirds of the true number of cases reaching at least the second trimester. The local register is much more complete, but time trends locally and nationally are similar. In England and Wales total prevalence declined from about 34 per 10000 live and stillbirths in 1974 to a plateau of just under 8 per 10000 in the 1990s. CONCLUSIONS: The decline in NTD prevalence is real and seems to have stopped. How this relates to changes in diet or the practice of vitamin supplementation is unknown, and the implications of the plateau are uncertain. OPCS figures of 500 NTDs annually in England and Wales represent about two thirds of the true number of cases.

England↗
PubMed1996

Transmit aperture processing for nonlinear contrast agent imaging.

Nonlinear contrast agents, such as bubbles, are used in ultrasound to enhance backscatter from blood. To increase contrast between these agents and tissue, nonlinear imaging methods, such as harmonic imaging or difference frequency imaging, can be used. For these, power is transmitted at one frequency and received at a different frequency. Nonlinear imaging methods, however, suffer from reduced contrast because of signal transmitted by the array at the receive frequency that linearly reflects off tissue. In this paper, transmit aperture processing is proposed to improve contrast in nonlinear imaging by suppressing signal transmission at the desired receive frequency. A specific method, known as alternate phasing, is presented as an example of this approach. Simulation results show that alternate phasing greatly improves contrast between tissue and bubbles.

Blood Flow Velocity↗
PubMed1996

Strain imaging of coronary arteries with intraluminal ultrasound: experiments on an inhomogeneous phantom.

In coronary arteries, knowing the relative stiffness of atherosclerotic lesions can help physicians select the most appropriate therapeutic modality. Because soft material supports larger strains than hard, measurements of this quantity can distinguish tissue of differing stiffness. In a previous paper, we described techniques for computing displacements and strains in coronary arteries using an integrated angioplasty and imaging catheter. Here, we demonstrate that hard and soft materials in a tissue-mimicking phantom can be differentiated with this device. Because tissue motion cannot be distinguished from catheter motion a priori, we perform all computations in the coordinate system centered at the balloon's geometric center. This reference frame depends only on balloon shape and is independent of catheter motion. A specialized correlation-based, phase-sensitive speckle tracking algorithm has been developed to compute strain. Maximum phantom displacement was about 25 microns, and the maximum radial, normal strain was about 1.5 percent.

Catheterization↗
PubMed1996

Comparison of cyclic and linear analogs of vasoactive intestinal peptide.

A series of i-->i + 4 side-chain to side-chain lactam analogs of vasoactive intestinal peptide has been prepared in order to study the effect of cyclization on biological activity. In vitro, on guinea pig tracheal smooth muscle and on human bronchial tissue, approximately half of the cyclic analogs showed increased potency and half were decreased over the linear analogs. Several cyclic compounds were between 10- and 20-fold more potent and one was 290-fold more potent than the linear species. In vivo, in guinea pigs, the cyclic compounds showed increased potency by up to 70-fold and significantly enhanced duration of action as compared to linear compounds.

Amino Acid Sequence↗
PubMed1996

Transplant medications.

This article discusses the medications used most commonly to keep the lung transplant patient free of rejection and infection. Medications used in the treatment of rejection are categorized by their use; therefore, the most common three- and four-drug regimens used for induction, maintenance, and treatment of rejection are discussed. The most commonly used antibacterial, antiviral, and antifungal drugs also are identified. Tables listing the characteristics of the medications are included to make it easier for the reader to quickly identify the indications, mechanisms of action, major adverse reactions, and nursing implications of these medications.

Anti-Bacterial Agents↗
PubMed1995

Escherichia coli expression vectors containing a protein kinase recognition motif, His6-tag and hemagglutinin epitope.

Escherichia coli expression vectors, based on the pET system, were constructed to allow fusion of a protein kinase (PK) recognition motif, a hemagglutinin (HG) epitope-tag and a His6-tag at the N-terminal portion of a protein of interest. The fusion proteins, that result from expression using these vectors, can be phosphorylated in vitro using cAMP-dependent PK, immunoprecipitated using monoclonal antibody against the HG-epitope, and can be rapidly purified using a Ni2+ column.

Amino Acid Sequence↗
PubMed1995

Protein structure. Why have six-fold symmetry?

Why do proteins proteins that encircle DNA have six-fold symmetry? One important factor may be the economy in protein mass with which DNA can be encircled by six globular subunits arranged in a ring.

Animals↗
PubMed1995

Structural and functional similarities of prokaryotic and eukaryotic DNA polymerase sliding clamps.

The remarkable processivity of cellular replicative DNA polymerases derive their tight grip to DNA from a ring-shaped protein that encircles DNA and tethers the polymerase to the chromosome. The crystal structures of prototypical 'sliding clamps' of prokaryotes (beta subunit) and eukaryotes (PCNA) are ring shaped proteins for encircling DNA. Although beta is a dimer and PCNA is a trimer, their structures are nearly superimposable. Even though they are not hexamers, the sliding clamps have a pseudo 6-fold symmetry resulting from three globular domains comprising each beta monomer and two domains comprising each PCNA monomer. These domains have the same chain fold and are nearly identical in three-dimensions. The amino acid sequences of 11 beta and 13 PCNA proteins from different organisms have been aligned and studied to gain further insight into the relation between the structure and function of these sliding clamps. Furthermore, a putative embryonic form of PCNA is the size of beta and thus may encircle DNA as a dimer like the prokaryotic clamps.

Amino Acid Sequence↗