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S Knapp

Publications and source records attributed to S Knapp.

At least 37 records · Page 2Linked to original sources
PubMed1998

Thermodynamic characterization of non-sequence-specific DNA-binding by the Sso7d protein from Sulfolobus solfataricus.

We used isothermal titration calorimetry and fluorescence spectroscopy to investigate the thermodynamics of non-sequence-specific DNA-binding by the Sso7d protein from the archaeon Sulfolobus solfataricus. We report the Sso7d-poly(dGdC) binding thermodynamics as a function of buffer composition (Tris-HCl or phosphate), temperature (15 to 45 degrees C), pH (7.1 to 8.0), osmotic stress and solvent (H2O/2H2O), and compare it to poly (dAdT) binding; and we have previously also reported the salt concentration dependence. Binding isotherms can be represented by the McGhee-von Hippel model for non-cooperative binding, with a binding site size of four to five DNA base-pairs and binding free energies in the range DeltaG degrees approximately -7 to DeltaG degrees approximately -10 kcal mol-1, depending on experimental conditions. The non-specific nature of the binding is reflected in similar thermodynamics for binding to poly(dAdT) and poly(dGdC). The native lysine methylation of Sso7d has only minor effects on the binding thermodynamics. Sso7d binding to poly(dGdC) is endothermic at 25 degrees C with a binding enthalpy DeltaH degrees approximately 10 kcal mol-1 in both phosphate and Tris-HCl buffers at pH 7.6, indicating that DeltaH degrees does not include large contributions from coupled buffer ionization equilibria at this pH. The binding enthalpy is temperature dependent with a measured heat capacity change DeltaCp degrees=-0.25(+/-0.01) kcal mol-1 K-1 and extrapolations of thermodynamic data indicate that the complex is heat stable with exothermic binding close to the growth temperature (75 to 80 degreesC) of S. solfataricus. Addition of neutral solutes (osmotic stress) has minor effects on DeltaG degrees and the exchange of H2O for 2H2O has only a small effect on DeltaH degrees, consistent with the inference that complex formation is not accompanied by net changes in surface hydration. Thus, other mechanisms for the heat capacity change must be found. The observed thermodynamics is discussed in relation to the nature of non-sequence-specific DNA-binding by proteins.

Archaeal Proteins↗
PubMed1998

Prognostic value of MIP-1 alpha, TGF-beta 2, sELAM-1, and sVCAM-1 in patients with gram-positive sepsis.

The aim of the present study was to evaluate the potential prognostic value of MIP-1 alpha, TGF-beta 2, sELAM-1, and sVCAM-1 in patients with gram-positive sepsis. Twenty-eight patients with gram-positive sepsis were compared to 11 patients with gram-negative sepsis and 15 healthy volunteers. Sepsis was defined by the criteria of Bone et al. (Crit. Care Med. 21, 5447-5463, 1993) and by isolation of at least two positive blood cultures with gram-positive/gram-negative bacteria. Plasma samples for determination of the immunological parameters were collected daily. Analysis of cytokines and adhesion molecules was performed on days 0 (day of sepsis criteria fulfillment), 4, and 7 (or 1 day before death). In the gram-positive group 10 of 28 patients died; in the gram-negative group 4 of 11 died. Only sELAM-1 plasma concentrations were found to be a useful early parameter in predicting patients' outcome in gram-positive sepsis. sELAM-1 concentrations at the onset of the study (day 0) were significantly higher in the nonsurviving patients than those in the survivors. MIP-1 alpha levels were significantly higher only on days 4 and 7. With regard to the measured plasma concentrations we believe that MIP-1 alpha is not a useful parameter for predicting patients' prognosis. The increase of sVCAM-1 might play a role in the pathogenesis of gram-positive sepsis; however, it could not be relied upon as an early prognostic parameter. The potential role of TGF-beta 2 in the development of gram-positive sepsis could not evaluated in the present study, whereas routine measurements of TGF-beta 2 offered no additional prognostic information.

Adult↗
PubMed1998

Two-dimensional crystallization of the chaperonin TF55 from the hyperthermophilic archaeon Sulfolobus solfataricus.

Oligomers of the chaperonin TF55 from Sulfolobus solfataricus have been successfully crystallized in two dimensions via their interaction with a phospholipid monolayer at the air/liquid interface. Oligomer orientation was dependent upon the lipid headgroup used. A neutral lipid monolayer gave rise to small paracrystalline areas of TF55 side views, whereas a negatively charged lipid monolayer resulted in large coherent crystalline areas of the chaperonin in an end-on orientation. These 2D crystals had p312 symmetry (a = b = 162 A, gamma = 60 degrees). Two-dimensional projection structures of the end-on arrays were produced by electron microscopy and image processing techniques. Under the conditions used to grow the crystals, the protein formed complexes of two stacked nine-subunit rings with threefold symmetry.

Archaeal Proteins↗
PubMed1998

Assessment of the proper depth of endotracheal tube placement with the Trachlight.

STUDY OBJECTIVE: To test the hypothesis that the correct depth of the endotracheal tube can be confirmed by transillumination method using the Trachlight device, which is a newly introduced lighted stylet for guided, blind tracheal intubation. DESIGN: Prospective, nonrandomized pilot-trial. SETTING: University-affiliated hospital. PATIENTS: 72 intubated and mechanically ventilated critically ill adult patients. INTERVENTIONS: Patients were briefly disconnected from the respirator. The centimeter marks of the Trachlight cather were brought into line with those of the endotracheal tube, resulting in congruence of the bulb of the Trachlight and the tube tip. MEASUREMENTS AND MAIN RESULTS: To measure the distance from the tip of the tube to that of the sternal notch, the Trachlight wand was retracted without moving the endotracheal tube until the bright light appeared in the sternal notch. A chest radiograph was taken to measure the distance between the carina and the tip of the tube. The Trachlight showed a distance of 4.0 +/- 1.3 cm from the tip of the tube to the sternal notch. Chest radiography revealed a distance of 3.3 +/- 1.6 cm between the carina and the tip of the tube, so that the calculated distance between sternal notch and carina was 7.3 +/- 1.5 cm. CONCLUSION: To achieve proper depth of the endotracheal tube, it is recommended that the tip of the endotracheal tube be placed 3 cm beyond the sternal notch. The Trachlight provides a simple and easy technique to achieve this goal.

Adult↗
PubMed1998

Architecture of nonspecific protein-DNA interactions in the Sso7d-DNA complex.

Many biochemical processes, including DNA packing, maintenance and control, rely on non-sequence specific protein-DNA interactions. Nonspecific DNA-binding proteins have evolved to tolerate a wide range of DNA sequences, yet bind with a respectable affinity. The nonspecific binding requirement is in contrast to that imposed on, for example, transcription factors and implies a different structural basis for the biomolecular recognition process. To address this issue, and the mechanism for archaeal DNA packing, we determined the structure of the Sso7d protein from Sulfolobus solfataricus in complex with DNA. Sso7d binds DNA by placing a triple-stranded beta-sheet across the DNA minor groove. The protein is anchored in this position by the insertion of hydrogen bond-donating side chains into the groove and additionally stabilized by electrostatic and non-polar interactions with the DNA backbone. This structure explains how strong binding can be achieved independent of DNA sequence. Sso7d binding also distorts the DNA conformation and introduces significant unwinding of the helix. This effect suggests a mechanism for DNA packing in Sulfolobus based on negative DNA supercoiling.

Amino Acid Sequence↗
PubMed1998

Time course of immunological markers in patients with the systemic inflammatory response syndrome: evaluation of sCD14, sVCAM-1, sELAM-1, MIP-1 alpha and TGF-beta 2.

BACKGROUND: The systemic inflammatory response syndrome (SIRS) is viewed as a system-wide inflammatory response. Up until now, no parameter has been available for predicting the development of septic shock. In the present study, we evaluated the usefulness of serum levels of CD14, vascular cells adhesion molecule-1 (VCAM-1), endothelial leucocyte adhesion molecule-1 (ELAM-1), macrophage inflammatory protein (MIP) 1 alpha and transforming growth factor beta 2 (TGF-beta 2) as early markers of outcome in patients with SIRS. METHODS: A group of 28 SIRS patients (13 survivors/15 non-survivors) was compared with a healthy control group and with patients with local inflammation. Blood samples were analysed on days 0, 4 and 7. Proinflammatory parameters such as sCD14, sVCAM-1, sELAM-1, MIP-1 alpha and anti-inflammatory parameters such as TGF-beta 2 were determined using enzyme-linked immunosorbent assay (ELISA). RESULTS: At the beginning, all evaluated proinflammatory immunological parameters with the exception of sVCAM-1 were significantly increased in patients with SIRS compared with the healthy control group. However, no significant difference could be observed for all immunological parameters comparing survivors and non-survivors, with the exception of interleukin (IL) 6 at day 7. CONCLUSION: All evaluated proinflammatory parameters were increased in patients with SIRS during the course of the disease. However, the parameters have no correlation with outcome and prognosis of SIRS patients.

Adult↗
PubMed1998

Comparison of oxygen cost of breathing with pressure-support ventilation and biphasic intermittent positive airway pressure ventilation.

OBJECTIVE: To assess the oxygen cost of breathing with either pressure-support ventilation (PSV) or biphasic intermittent positive airway pressure ventilation (BIPAP). DESIGN: Prospective, randomized, crossover study. SETTING: Medical intensive care unit of a university hospital. PATIENTS: Twenty clinically stable and spontaneously breathing patients after long-term mechanical ventilation. INTERVENTIONS: Patients were randomized to start on either PSV or BIPAP, and measurements were performed after an adaptation period of 30 mins. Immediately after, the ventilatory mode was changed and after another 30-min adaptation period, the same measurements were performed. MEASUREMENTS AND MAIN RESULTS: Indirect calorimetry was performed during each ventilatory mode for a period of 30 mins. Oxygen consumption, energy expenditure, CO2 production, and respiratory quotient did not differ significantly between the two ventilatory modes, regardless of the patients' randomization. There were no statistically significant differences with regard to respiratory rate, minute volume, and blood gas analysis. All patients tolerated both ventilatory modes without any signs of discomfort. CONCLUSIONS: Pressure support ventilation and BIPAP are both used for weaning patients gradually from the ventilator. BIPAP may be advantageous in patients not breathing sufficiently with PSV, since no patient effort is necessary with use of this ventilatory mode.

Adult↗
PubMed1998

Diagnostic validity of pulmonary artery catheterization for residents at an intensive care unit.

OBJECTIVE: To assess the amount of additional information provided by measurements derived from pulmonary artery catheter (PAC) use beyond that derived from clinical evaluation by intensive care residents. METHODS: One hundred forty-nine consecutive patients undergoing right-heart catheterization were prospectively included in the study. Before inserting a PAC, physicians had to predict pulmonary arterial pressure (PAP), pulmonary capillary wedge pressure (PCWP), systemic vascular resistance index (SVRI), cardiac index (CI), mixed venous oxygen saturation (SvO2), oxygen delivery (DO2), oxygen consumption (VO2), and pulmonary shunt fraction (Qs/Qt) by selecting a given option on a questionnaire. Ranges of options were chosen to create clear clinical differences among them. RESULTS: The correct value was predicted in a median of 50% of cases (range, 45-63%). PAP was predicted correctly in 55%, PCWP in 46%, SVRI in 63%, CI in 62%, SvO2 in 45%, DO2 in 45%, VO2 in 51%, and Qs/Qt in 51%. A significant difference was found between estimated and measured values for all parameters (p < 0.01). No significant differences were detected between more and less experienced physicians. There was no significant difference between estimated and measured values with respect to the different courses of intensive care unit admissions or the different indications for PAC insertion. CONCLUSION: In a selected group of critically ill patients, the PAC adds valuable and clinically relevant information to clinical assessment in about 50% of cases. Its use should not be withheld in patients with unclear hemodynamic and metabolic profiles.

Catheterization, Swan-Ganz↗
PubMed1998

[Experiences with inhalative nitric oxide in internal medicine intensive care patients with adult respiratory distress syndrome].

Inhalative nitric oxide (NO) has recently been included in the therapeutic armament for the treatment of ARDS. We evaluated the effect of inhalative NO on hemodynamic and oxygen transport parameters in 30 internal intensive care patients suffering from ARDS. All patients received a pulmonary artery catheter. Hemodynamics were assessed prior to NO therapy and after 1, 6, 12, and 24 h. 80% (n = 24) of the patients were classified as therapy responders. The median NO dose was 15 ppm (range 5 to 40 ppm). The PaO2/FiO2--ratio increased significantly after initiation of NO (p = 0.0002) while the pulmonary shunt fraction (Qs/Qt) decreased significantly (p = 0.0019). All other measured or calculated parameters including arterial and pulmonary arterial blood pressure remained unchanged. No negative effects of the therapy could be observed. Inhalative NO improves oxygenation in most intensive care patients with ARDS and thus offers the possibility to reduce invasiveness of mechanical ventilation.

Administration, Inhalation↗
PubMed1997

Renal tolerability of four different once-daily dose regimen of netilmicin in critical care patients.

A prospective, randomized trial was conducted in a medical intensive care unit to assess safety and tolerability of four different dose regimens of intravenous netilmicin given once daily in the treatment of febrile episodes in critically ill patients. Eighty patients with febrile episodes during their stay in the intensive care unit were included in the study. The patients were randomized into four groups: Group 1 received a single daily dose of netilmicin based upon weight, age and renal function according to a dosage nomogram [13] (mean dose 298 +/- 29 mg, median 300 mg, range 250-350 mg), group 2 received 150% of this standard dose (mean 418 +/- 45 mg, median 400 mg, range 350-500 mg), group 3 200% (mean 525 +/- 41 mg, median 500 mg, range 400-550 mg) and group 4 250% (mean 710 +/- 39 mg, median 650 mg, range 600-750 mg). Duration of treatment was six days. Positive cultures were obtained in 29 patients. Serum creatinine and creatinine clearance, as well as netilmicin trough levels and levels of alpha 1-microglobulin showed no significant difference between the groups before, during, and after therapy. Our results indicate that with once daily dosing even high doses of netilmicin are well tolerated in patients with a creatinine clearance of > 70 ml/min before therapy. Necessary precautions include monitoring of drug trough levels (< 1 mg/L) and maintenance of adequate volume status.

Acute Kidney Injury↗
PubMed1997

Crystal structure of glutamate dehydrogenase from the hyperthermophilic eubacterium Thermotoga maritima at 3.0 A resolution.

The extremely thermostable glutamate dehydrogenase from the hyperthermophilic bacterium Thermotoga maritima has been crystallized and the three-dimensional structure has been determined by X-ray diffraction methods. Crystals up to a maximum size of 1.2 mm have been grown in 3% polyethylene glycol, 120 mM ammonium acetate and 50 mM bis-tris propane (pH 6.5). The enzyme crystallized in the trigonal space group P3(1)21 with the cell dimensions a = b = 147.3 A, c = 273.6 A. The diffraction limit of these crystals is 3.0 A. Measured diffraction data have a completeness of 94% up to a resolution of 3.0 A and contain 75% of all possible data in the last resolution shell between 3.1 and 3.0 A. The crystal structure of T. maritima glutamate dehydrogenase has been solved by Patterson search methods using the hexameric Pyrococcus furiosus glutamate dehydrogenase as a search model. The crystallographic refinement has been carried out to a maximum resolution of 3.1 A and an crystallographic R-value of 22.5% (Rfree = 29.5%). The three-dimensional structure of the T. maritima enzyme shows typical features of hexameric glutamate dehydrogenases: six subunits are arranged in 32 symmetry. Each subunit consists of two domains connected by a flexible hinge region. Secondary structure elements as well as residues important for the catalytic activity of the enzyme are highly conserved. A structural comparison of the two glutamate dehydrogenases from the hyperthermophiles T. maritima and P. furiosus with the enzyme from the mesophilic bacterium Clostridium symbiosum has revealed that common as well as distinct mechanisms contribute to the thermal stability of these enzymes. The number of intrasubunit ion pairs is increased and the volume of intrasubunit cavities decreased in both thermostable enzymes, whereas striking differences have been observed in the subunit interfaces. In P. furiosus glutamate dehydrogenase the subunit interactions are dominated by ionic interactions realized by large saltbridge networks. However, in T. maritima glutamate dehydrogenase the number of intersubunit ion pairs is reduced and the hydrophobic interactions are increased.

Amino Acid Sequence↗
PubMed1997

Diversity of the immunodominant epitope of gp41 of HIV-1 subtype O and its validity for antibody detection.

The immunodominant regions of the gp41 from 13 HIV-1 subtype O strains from Cameroon, 11 from France and one from Germany were sequenced. The amino acid sequences were compared to those of the 3 published HIV-1 subtype O isolates, ANT70, MVP-5180 and VAU. All HIV-1 subtype O isolates had a very conserved amino acid sequence in this region and showed a subtype O specific structure. Within the cysteine loop there was a positive charge of two basic amino acids, arginine and lysine. Only two strains (CM.6778 and CM.8161) showed an acidic amino acid in this loop. None of the isolates showed the same amino acid sequence in this immunodominant region. A 25 residue peptide from the immunodominant domain of gp41 of the MVP-5180 strain was synthesized, cycled to form the cysteine-loop and coated to microtiter plates. Antibody binding was detected by indirect ELISA using an enzyme labeled anti-human IgG. Out of 111 anti-HIV-1 positive specimens, collected mainly from Cameroonian HIV infected patients, only 10 were not reactive in this assay. The 42 anti-HIV-1 subtype O positive specimens gave all a reaction above cut off. Despite the diversity found in the amino acid sequences within the 25 isolates a peptide-based indirect ELISA representing the immunodominant epitope of the strain MVP-5180 successfully detected all the anti-HIV-O sera so far tested, pointing to the importance of adding such a peptide for correct identification of HIV-1 subtype O infected patients, while some assays without HIV-O specific antigens partially fail to detect all anti-HIV-O specimens.

Amino Acid Sequence↗
PubMed1997

Exogenous surfactant therapy in a patient with adult respiratory distress syndrome after near drowning.

A 24-year-old woman developed adult respiratory distress syndrome (ARDS) after near-drowning due to attempted suicide. Conventional mechanical ventilation together with prone positioning and inhaled nitric oxide could not provide sufficient oxygenation. Surface tension data (gamma min = 27 dyn/cm, stability index = 0.341) from a lavage sample supported the hypothesis that the surfactant function of this patient was drastically reduced due to a washout effect by aspiration of fresh water. Porcine surfactant (Curosurf, 50 mg/kg for each lung) was instilled via fibreoptic bronchoscope. The partial arterial carbon dioxide pressure (paCO2) and fraction of inspired oxygen (FiO2) ratio as well as shunt fraction (Qs/Qt) improved impressively. When respiratory situation deteriorated again, surfactant application was repeated. Altogether, six bolus instillations of surfactant (total dose 300 mg/kg = 18,000 mg) were administered until the respiratory situation had stabilized and oxygenation could be maintained by conventional mechanical ventilation. The radiological findings did not show substantial amelioration. The patient developed septic shock and died 12 days after admission. Surfactant application apparently led to a significant improvement of the respiratory function. However, the outcome could not be influenced positively. The high cost of surfactant therapy prevents the more widespread early administration in patients at risk.

Adult↗
PubMed1997

Prostaglandin E1 inhibits platelet decrease after massive blood transfusions during major surgery: influence on coagulation cascade?

BACKGROUND: A plunge in platelet count if often observed after massive blood transfusions during major surgery. This study was designed to assess whether the prophylactic administration of prostaglandin E1 (PGE1) might prevent this drop in platelet count. METHODS: Forty-five patients receiving massive transfusions of packed red blood cells (> 10 units) during major orthopedic surgery were enrolled in a prospective, randomized, double-blind, placebo-controlled study and divided into two groups: group 1 (therapy group) received intravenous PGE1 up to 30 ng/kg/min for 72 hours after surgery, and group 2 (placebo group) received a placebo during the same time period. RESULTS: The patients in group 1 suffered no reduction in platelet count and thus required no platelet concentrate transfusions. In contrast, a significant postoperative drop in platelet count (p < 0.05) was observed in the placebo group between days 3 and 5 after surgery when compared to the therapy group, necessitating transfusions of platelet concentrates in this group. Similarly, red blood cell count and hemoglobin were far more stable in the therapy group, which required fewer transfusions of red blood cells than did the placebo group (p < 0.05). There seemed to be a tendency toward a consumptive coagulation disorder in the placebo group as indicated by a decrease of fibrinogen levels, prothrombin time, and antithrombin III activity, and an increase of partial thromboplastin time. The incidence of adult respiratory distress syndrome was slightly lower in the therapy group. Last but not least, the length of intensive care unit stay was significantly shorter in the therapy group (p < 0.05). CONCLUSION: In our study, the administration of PGE1 prevented a reduction in platelet count. Furthermore, measurements of clotting activity furnished the possibility that PGE1 might inhibit transfusion-induced coagulation disturbances. We recommend that PGE1 should be considered in patients requiring massive transfusion during major surgery.

Adult↗
PubMed1997

Azygos arch cannulation by central venous catheters: radiographic detection of malposition and subsequent complications.

The aim of this study was to determine the frequency of radiographically evident central venous catheter misplacement in the azygos arch and to analyze whether the frequency of azygos arch cannulation is dependent on the anatomical site of catheter insertion. We reviewed 1,287 postprocedural examinations and 3,441 follow-up examinations. Catheters had been inserted through the left (6%) or right (15%) internal jugular veins and through the left (32%) or right (46%) subclavin veins. Radiographs were analyzed for possible catheter malposition in the azygos arch and for complications related to this malposition. Catheter malposition in the azygos arch was seen on 16/1,287 (1.2%) postprocedural radiographic examinations. Of the 16 malpositioned catheters, 11 (69%) had been inserted in the left subclavian vein, three (19%) in the left jugular vein, two (12%) in the right subclavian vein, and none (0%) in the right jugular vein. There was a statistically significant difference in the frequency of azygos arch cannulation between left- and right-sided catheters (p = 0.001). All complications consisted of venous perforations and were seen in three of 16 cases (19%). Azygos arch cannulation is a rare but hazardous central venous catheter malposition that occurs early after catheter insertion and carries a substantial risk for complication. The risk for azygos arch cannulation is substantially increased if catheters are inserted in left-sided veins. Because of the severity of subsequent complications, radiologists should be vigilant in the detection of this rare malposition.

Azygos Vein↗
PubMed1996

Thermal unfolding of the DNA-binding protein Sso7d from the hyperthermophile Sulfolobus solfataricus.

Thermal unfolding of the small hyperthermophilic DNA-binding protein Sso7d was studied by circular dichroism spectroscopy and differential scanning calorimetry. The unfolding transition can be described by a reversible two state process. Maximum stability was observed in the region between pH 4.5 and 7.0 where Sso7d unfolds with a melting temperature between 370.8 to 371.9 K and an unfolding enthalpy between 62.9 and 65.4 kcal/mol. The heat capacity differences between the native and the heat denatured states obtained by differential scanning calorimetry (620 cal/(molK)) and circular dichroism spectroscopy (580 cal/(mol K)) resulted in comparable values. The thermodynamic reason for the high melting temperature of Sso7d is the shallow stability curve with a broad free energy maximum, corresponding to the relatively small heat capacity change which was obtained. The calculated stability curve shows that Sso7d has, despite of its high melting temperature, an only moderate intrinsic stability, which reaches its maximum (approximately 7 kcal/mol) at 282 K. Sso7d is particularly poorly stabilized (approximately 1 kcal/mol) at the maximum physiological growth temperature of Sulfolobus solfataricus. Sso7d has furthermore untypically low specific enthalpy (0.99 kcal/(mol residue)) and entropy (2.99 cal/(mol K)) values at convergence temperatures. No significant differences in thermal stability of the partially methylated Sso7d from Sulfolobus solfataricus and the cloned non-methylated form of the protein expressed in Escherichia coli were observed.

Amino Acid Sequence↗