Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “CLAMPS”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,639 records · Page 91Linked to original sources
PubMed1995

Experimental delayed postischemic spinal cord hypoperfusion after aortic cross-clamping.

BACKGROUND: As in the brain, recent evidence has suggested a defect in the microcirculation during the reperfusion period after spinal cord ischemia. This investigation was undertaken in order to delineate blood flow dynamics in the postischemic spinal cord of the rat. METHODS: Male Sprague-Dawley rats underwent cross-clamping of the aorta and subclavian arteries (XC) for 11 minutes. Spinal cord blood flow (SCBF) was measured by autoradiography in the gray and white matter of cervical (Ce), thoracic (Th) and lumbar (Lu) regions during XC, 1 h, 6 h and 24 h (XC n = 8, 1 h n = 9, 6 h n = 9, and 24 h n = 11, groups) after XC. Control groups underwent surgical manipulations and SCBF measurement but no XC (Sham 1, n = 8), or clamping of the subclavian arteries only (Sham 2, n = 8). RESULTS: In Ce cord, there was no difference between SCBF of 1 h, 6 h, 24 h and Sham 1. In Th cord, SCBF was reduced during XC (P < 0.003 vs. Sham 2), 1 h, 6 h (P < 0.04 and P < 0.01 vs. Sham 1). In Lu cord, SCBF was not detectable in XC, and depressed in 1 h (P < 0.003) and 6 h (P < 0.003). There was no difference between 24 h and Sham 1 in Ce, Th, and Lu cords. CONCLUSIONS: The study demonstrated a period of delayed postischemic hypoperfusion in the white and gray matter of Th and Lu cord segments lasting 6 h after XC. The phenomenon may play an important role in the ultimate fate of neural elements with borderline viability after ischemic injury.

Animals↗
PubMed2004

Inhibition of protein interactions with the beta 2 sliding clamp of Escherichia coli DNA polymerase III by peptides from beta 2-binding proteins.

The sliding clamp of the Escherichia coli replisome is now understood to interact with many proteins involved in DNA synthesis and repair. A universal interaction motif is proposed to be one mechanism by which those proteins bind the E. coli sliding clamp, a homodimer of the beta subunit, at a single site on the dimer. The numerous beta(2)-binding proteins have various versions of the consensus interaction motif, including a related hexameric sequence. To determine if the variants of the motif could contribute to the competition of the beta-binding proteins for the beta(2) site, synthetic peptides derived from the putative beta(2)-binding motifs were assessed for their abilities to inhibit protein-beta(2) interactions, to bind directly to beta(2), and to inhibit DNA synthesis in vitro. A hierarchy emerged, which was consistent with sequence similarity to the pentameric consensus motif, QL(S/D)LF, and peptides containing proposed hexameric motifs were shown to have activities comparable to those containing the consensus sequence. The hierarchy of peptide binding may be indicative of a competitive hierarchy for the binding of proteins to beta(2) in various stages or circumstances of DNA replication and repair.

Amino Acid Motifs↗
PubMed1997

Dissociation of bacteriophage T4 DNA polymerase and its processivity clamp after completion of Okazaki fragment synthesis.

The mechanism of bacteriophage T4 DNA polymerase (gp43) and clamp (gp45) protein dissociation from the holoenzyme DNA complex was investigated under conditions simulating the environment encountered upon completion of an Okazaki fragment. Lagging strand DNA synthesis was approximated using a synthetic construct comprised of a doubly biotinylated, streptavidin-bound 62-mer DNA template, paired with complementary primers to generate an internal 12-base gap where the 5'-end primer contained either a 5'-OH (DNA primer) or a 5'-triphosphate (RNA primer) group. Rapid kinetic measurements revealed that upon encountering the blocking primer, the holoenzyme either dissociates from DNA (approximately 40%) or strand-displaces the blocking strand (approximately 60%). The two blocking oligonucleotides (DNA or RNA) induce a 30-50-fold increase in the rate of holoenzyme dissociation, with both polymerase and clamp proteins dissociating simultaneously. Inhibition of ATP hydrolysis by ATP-gamma-S did not have a measurable effect upon holoenzyme dissociation from DNA. The presence of gp32, the single-strand binding protein, caused a small (3-fold) increase in the rate constant for dissociation.

Bacteriophage T4↗
PubMed2000

Tracking sliding clamp opening and closing during bacteriophage T4 DNA polymerase holoenzyme assembly.

The bacteriophage T4 DNA polymerase holoenzyme, consisting of the DNA polymerase (gp43), the sliding clamp (gp45), and the clamp loader (gp44/62), is loaded onto DNA in an ATP-dependent, multistep reaction. The trimeric, ring-shaped gp45 is loaded onto DNA such that the DNA passes through the center of the ring. gp43 binds to this complex, thereby forming a topological link with the DNA and increasing its processivity. Using stopped-flow fluorescence-resonance energy transfer, we have investigated opening and closing of the gp45 ring during the holoenzyme assembly process. Two amino acids that lie on opposite sides of the gp45 subunit interface, W91 and V162C labeled with coumarin, were used as the fluorescence donor and acceptor, respectively. Free in solution, gp45 has two closed subunit interfaces with W91 to V162-coumarin distances of 19 A and one open subunit interface with a W91 to V162C-coumarin distance of 40 A. Making the assumption that the distance across the two closed subunit interfaces is unchanged during the holoenzyme assembly process, we have found that the distance across the open subunit interface is first increased to greater than 45 A and is then decreased to 30 A during a 10-step assembly mechanism. The gp45 ring is not completely closed in the holoenzyme complex, consistent with previous evidence suggesting that the C-terminus of gp43 is inserted into the gp45 subunit interface. Unexpectedly, ATP-hydrolysis events are coupled to only a fraction of the total distance change, with conformational changes linked to binding DNA and gp43 coupled to the majority of the total distance change. Using the nonhydrolyzable ATP analogue ATP-gamma-S results in formation of a nonproductive gp45 x gp44/62 complex; however, adding an excess of ATP to this nonproductive complex results in rapid ATP/ATP-gamma-S exchange to yield a productive gp45 x gp44/62 complex within seconds.

Adenosine Diphosphate↗
PubMed2006

A third mode of DNA binding: Phosphate clamps by a polynuclear platinum complex.

We describe a 1.2 A X-ray structure of a double-stranded B-DNA dodecamer (the Dickerson Dodecamer, DDD, [d(CGCGAATTCGCG)]2) associated with a cytotoxic platinum(II) complex, [{trans-Pt(NH3)2(NH2(CH2)6(NH3+)}2-mu-{trans-Pt(NH3)2(NH2(CH2)6NH2)2}] (TriplatinNC). TriplatinNC is a multifunctional DNA ligand, with three cationic Pt(II) centers, and directional hydrogen bonding functionalities, linked by flexible hydrophobic segments, but without the potential for covalent interaction. TriplatinNC does not intercalate nor does it bind in either groove. Instead, it binds to phosphate oxygen atoms and thus associates with the backbone. The three square-planar tetra-am(m)ine Pt(II) coordination units form bidentate N...O...N complexes with OP atoms, in a motif we call the Phosphate Clamp. The geometry is conserved among the 8 observed phosphate clamps in this structure. The interaction appears to prefer O2P over O1P atoms (frequency of interaction is O2P > O1P, base and sugar oxygens > N). The high repetition and geometric regularity of the motif suggests that this type of Pt(II) center can be developed as a modular nucleic acid binding device with general utility. TriplatinNC extends along the phosphate backbone, in a mode of binding we call "Backbone Tracking" and spans the minor groove in a mode of binding we call "Groove Spanning". Electrostatic forces appear to induce modest DNA bending into the major groove. This bending may be related to the direct coordination of a sodium cation by a DNA base, with unprecedented inner-shell (direct) coordination of penta-hydrated sodium at the O6 atom of a guanine.

Amines↗
PubMed2000

Cardiac output measurements during cross-clamping of the descending thoracic aorta in pigs. A comparison between transit-time ultrasound, thermodilution and pulsed Doppler ultrasound.

BACKGROUND: Cross-clamping of the descending thoracic aorta (XC) induces an increase in cardiac output (CO). The intention of this study was to evaluate the high CO during XC by the use of clinically available methods (thermodilution and pulsed Doppler ultrasound) compared to transit-time ultrasound flowmetry of the ascending aorta as the gold standard. METHOD: Ten pigs were anaesthetised with ketamine and fentanyl. The descending thoracic aorta was cross-clamped for 30 min, and cardiac output was measured with pulmonary artery thermodilution technique, pulsed Doppler ultrasound on the aortic annulus and transit-time ultrasound flowmetry of the ascending aorta. RESULTS: At 15 min following XC, CO increased from 1.7 l/min to 4.6 l/min measured with transit-time ultrasound (P<0.05). With thermodilution technique, CO increased from 2.6ll/min to 5.7 l/min (P<0.05), and from 2.4 l/min to 6.0 l/min measured with Doppler ultrasound (P<0.05). There was an increase in mean arterial pressure of 81% and heart rate increased 76% (P<0.05). CONCLUSION: XC of the descending thoracic aorta induces an increase in CO of 171%. Thermodilution and pulsed Doppler ultrasound are reliable methods for detecting high cardiac output during thoracic aortic surgery.

Animals↗
PubMedDate not supplied

K channels in T lymphocytes: a patch clamp study using monoclonal antibody adhesion.

Ionic fluxes are thought to be involved in mediating the proliferation of peripheral blood lymphocytes (PBLs) in response to mitogenic substances. Among the earliest events occurring after the addition of mitogen to cultured lymphocytes are changes in rates of cation transport. We were interested, therefore, in the possible role of ion channels in mediating the lymphocyte proliferative response. The development of patch clamp techniques by Neher and colleagues has made it possible to study membrane conductances in a variety of small cell types. We have developed a method which uses monoclonal antibodies to make cells adhere to solid surfaces for two major reasons: (1) it is much easier to patch clamp stationary cells, and (2) the method can be used to selectively adhere a particular cell type from a heterogeneous population. We have used these techniques here to identify whole-cell potassium currents, in lymphocytes recognized by OKT11 monoclonal antibody, which are increased 1.9-fold by mitogenic stimulation.

Antibodies, Monoclonal↗
PubMedDate not supplied

Patch-clamping of the inner mitochondrial membrane reveals a voltage-dependent ion channel.

The prime function of mitochondria is to provide the cell with adenosine triphosphate (ATP). ATP synthesis is driven by the protonmotive force (delta p), which is generated and maintained across the inner mitochondrial membrane (IMM) by the activity of the respiratory chain. It is widely believed that the IMM is unlikely to contain ion channels like those present in the plasma membrane, because the high rates of ion transport characteristic of open channels would be expected to dissipate the delta p. Although the small size of the organelle has prevented the use of classical electrophysiological methods, the recent introduction of the patch-clamp technique, which allows currents to be recorded from very small cells, has enabled us to test this hypothesis. By patch-clamping the IMM, we have identified a slightly anion-selective channel, which is voltage-dependent and has a mean conductance of 107 pS in the presence of symmetrical 150 mM KCl.

Adenosine Triphosphate↗
PubMed1990

Neuropeptide modulation of single calcium and potassium channels detected with a new patch clamp configuration.

Calcium channel activity is crucial for secretion and synaptic transmission, but it has been difficult to study Ca2+ channel modulation because survival and regulation of some of these channels require cytoplasmic constituents that are lost with the formation of cell-free patches. Here we report a new patch clamp configuration in which activity and regulation of channels are maintained after removal from cells. A pipette containing the pore-forming agent nystatin is sealed onto a cell and withdrawn to form an enclosed vesicle. The resulting perforated vesicle, formed from pituitary tumour cells, contains Ca2+ and K+ channels. Ca2(+)-activated K+ channels in the vesicle are activated by cyclic AMP analogues, and by a neuropeptide (thyrotropin-releasing hormone) that stimulates phosphatidylinositol turnover and inositol trisphosphate-gated Ca2+ release from intracellular organelles. Thus, the perforated vesicle retains signal transduction systems necessary for ion channel modulation. Functional dihydropyridine-sensitive Ca2+ channels (L-type) are maintained in the vesicle, and their gating is inhibited by thyrotropin-releasing hormone. Hence, this new patch clamp configuration has allowed a direct detection of the single-channel basis of transmitter-induced inhibition of L-type Ca2+ channels. The modulation of Ca2(+)-channel gating may be an important mechanism for regulating hormone secretion from pituitary cells.

Calcium↗
PubMed2006

Communication between subunits within an archaeal clamp-loader complex.

We have investigated the communication between subunits in replication factor C (RFC) from Archaeoglobus fulgidus. Mutation of the proposed arginine finger in the small subunits results in a complex that can still bind ATP but has impaired clamp-loading activity, a process that normally only requires binding of nucleotide. The small subunit alone forms a hexameric ring that is six-fold symmetric in the absence of ATP. However, this symmetry is broken when the nucleotide is bound to the complex. A conformational change associated with nucleotide binding may relate to the opening of PCNA rings by RFC during the loading reaction. The structures also reveal the importance of the N-terminal helix of each subunit at the ATP-binding site. Analysis of mutant protein complexes containing subunits lacking this N-terminal helix reveals key distinct regulatory roles during clamp loading that are different for the large and small subunits in the RFC complex.

Adenosine Triphosphate↗
PubMed2002

PCNA clamp facilitates action of DNA cytosine methyltransferase 1 on hemimethylated DNA.

BACKGROUND: Proliferating cell nuclear antigen (PCNA) is a ring-shaped protein known as a processivity factor of DNA polymerase delta. In addition to this role, PCNA interacts with a number of other proteins to increase their local concentration at replicated DNA sites. DNA cytosine methyltransferase 1 (Dnmt1), which preserves epigenetic signals by completing the methylation of hemimethylated DNA after DNA replication, has been indicated as one of these PCNA binding proteins by a previous work. However, the molecular mechanisms and functional significance of their association have not yet been studied. RESULTS: Dnmt1 can be readily isolated from nuclear extracts by PCNA affinity chromatography. Studies of the interactions between the two proteins demonstrate that the N-terminal region of Dnmt1, which contains a typical PCNA binding motif, has core PCNA binding activity, and that the remaining portion of the protein exerts a negative influence on the interaction of Dnmt1 with PCNA. The affinity of Dnmt1 for DNA is much higher for DNA bound by PCNA than for free DNA. Furthermore, DNA methylation assays with hemimethylated DNA as a substrate revealed that PCNA clamp-bound DNA is methylated more efficiently by Dnmt1 than is free DNA. CONCLUSION: These results provide the first biochemical evidence that physical interactions between PCNA and Dnmt1 facilitate the methylation of newly neplicated DNA, on which PCNA remains associated as a functional clamp.

Animals↗
PubMed1983

A method for quantifying insulin sensitivity in vivo in the anesthetized rat: the euglycemic insulin clamp technique coupled with isotopic measurement of glucose turnover.

The euglycemic insulin clamp technique coupled with isotopic measurement of the glucose utilization rate ([3-3H] glucose), described previously in man, has been adapted to characterize and quantify insulin sensitivity in vivo in the rat. Only 30 min were needed for the exogenous glucose infusion rate, the specific activity of [3-3H] glucose and the plasma insulin to reach a steady-state value. During the next 30 min of the experiment, blood glucose was maintained at a constant basal level with a coefficient of variation of 2.3%. Plasma insulin was 48 +/- 4 microU/ml and glucose utilization was 1.84 +/- 0.07 mg/min in anesthetized female rats in the basal state. During euglycemic insulin clamp, plasma insulin was 302 +/- 24 microU/ml, glucose utilisation was increased by 53 +/- 10% and endogenous glucose production was decreased by 76 +/- 3%. By studying several insulin concentrations in different groups of rats, we hope to be able to characterize insulin resistance during pregnancy and to determine whether hepatic or peripheral tissues are responsible for this insulin resistance.

Animals↗
PubMed2001

A pair of clamps for a safe removing and repositioning of beta and gamma angular settings of Brown-Roberts-Wells stereotactic system during operation.

The Brown-Roberts-Wells arc system is a non-target-centered design, i.e., without an independent approach angle. The approach angle of this system strictly depends on precalculated values (entry and target point). Therefore, some components of the system used sometimes prevent a direct insight into the operation field. Once the entry point has been set, the arc system normally has to be taken off to permit an unimpeded approach to the burr hole. To facilitate rotation and return to the primary beta and gamma angular settings during stereotactic craniotomy and other surgery, a pair of clamps was designed for the BRW arc system. These clamps help the approach to the entry point in such a way that some components of the arc (e.g., the guide block holder) are removed from the surgical field, thus giving wide visual access for the stereotactic approach. Consequently, it is no longer necessary to remove the entire arc system, resulting in an increased operation safety and shorter operation times.

Equipment Design↗
PubMed2004

Intermittent aortic cross-clamping for isolated CABG can save lives and money: experience with 15307 patients.

BACKGROUND: The ideal myocardial protection during isolated CABG is still a matter of debate. Cardioplegia versus intermittent aortic cross-clamping (IACC) are the main opponents; the following article shows that IACC can be safe, efficient and might be cheaper than cardioplegia. METHODS: Demographics and co-morbidities of 15307 CABG only patients consecutively operated on between January 1993 and October 2001 in the Heart Center in Bad Oeynhausen were assessed by the German Quality Assurance data set and risk-stratified using the EuroSCORE. Outcome (30-day or in-hospital mortality) was compared to the expected EuroSCORE estimation. RESULTS: Expected mortality was 3.25 %, observed mortality was 1.3 %, being significantly lower in the low, medium as well as high risk patients subgroup. Complication rates increased steadily with expected mortality rates. Stroke and myocardial infarction rates for patients with peripheral vessel disease were not higher than in comparable studies. More than 1000000 EUR were saved by lower cardioplegia bills. CONCLUSION: Myocardial protection with intermittent aortic cross-clamping for isolated CABG can be safe, effective, and economically advantageous when compared to cardioplegic solutions.

Cardiopulmonary Bypass↗
PubMed1983

Clinical and quantitative birefringence assessment of 100 patients with aortic clamping periods in excess of 120 minutes after hypothermic cardioplegic arrest.

At St. Thomas' Hospital the first 100 patients with prolonged aortic cross-clamp times in excess of 120 minutes have been analyzed clinically (low cardiac output and mortality) and 49 of these patients from which left and right ventricular biopsies were taken, have been analyzed by quantitative birefringence (biophysical measurement of myocardial deterioration). A total of 8 patients died (8%) and 11 had low cardiac output syndrome (11%). The patients were divided into those given only a single infusion (n = 18) and those given hourly infusions (n = 82) of hypothermic cardioplegic solution. These were then subdivided into those with single (n = 37) or multiple (n = 63) corrective surgical procedures. In the patients who had a single corrective surgical procedure there was no difference at all, but in those patients who had multiple corrective surgical procedures, hourly infusion reduced low cardiac output from 12.5% to 9.4% in multiple valve patients and from 50% to 19% in bypass graft combined with valve replacement patients. In this latter group mortality fell from 50% to 4.8% but there were only 2 patients given a single infusion. There was no statistically significant advantage in hourly infusions compared with single infusions, either clinically or cytochemically. Twenty-six patients had aortic cross-clamp periods in excess of 150 minutes. Mortality and low cardiac output increased compared with the 120 to 150 minute group, rising from 3% to 19% and from 7% to 23% respectively. As a result of these analyses, surgical practice has been changed to 30-minute reinfusion intervals with currently improved results.

Birefringence↗
PubMed1988

[Hypoglycemic and hyperinsulinemic clamp technic to determine the secretory behavior of insulinoma].

In two female patients of 62 and 76 years of age, respectively, who had insulinomas subsequently confirmed by histology, the secretion pattern of insulin-producing tumours was studied. In the 62-year old patient complete reproducible suppression of insulinoma secretion was achieved by means of large exogenous doses of insulin, whereas this was not possible in the 76-year old patient. A modified hypoglycaemic clamping technique was used in the study. It is concluded that, depending on plasma insulin levels, relative suppression of insulinoma is possible, and it is recommended to use the described differentiated clamping procedure to investigate the secretion pattern of autonomous insulin-producing tumours.

Adenoma, Islet Cell↗
PubMed2006

Protein folding in a force clamp.

Kinetics of folding of a protein held in a force clamp are compared to an unconstrained folding. The comparison is made within a simple topology-based dynamical model of ubiquitin. We demonstrate that the experimentally observed variations in the end-to-end distance reflect microscopic events during folding. However, the folding scenarios in and out of the force clamp are distinct.

Protein Folding↗