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C Block

Publications and source records attributed to C Block.

At least 19 recordsLinked to original sources
PubMed2001

Domain shuffling as a tool for investigation of protein function: substitution of the cysteine-rich region of Raf kinase and PKC eta for that of yeast Pkc1p.

With the completion of the sequences of entire genomes, the need for functional characterisation of proteins and their domains is becoming acute. Conserved regions within proteins often share overlapping functions but despite this conservation may fulfil quite different tasks in different species. In this work, we investigated the cysteine-rich motif (C1 domain) of yeast protein kinase C (Pkc1p) as a model to establish a test system for domain function. C1 domains activate kinases through binding of either diacylglycerol and/or phosphatidylserine, as in many members of the protein kinase C (PKC) family, or by binding small GTPases, as in Raf kinase. In contrast to other members of the protein kinase C superfamily, Pkc1p of Saccharomyces cerevisiae is activated via binding of the small G-protein Rho1p to its C1 domain. We developed a system for domain shuffling to establish the function of C1 domains from human Raf kinase and rat PKC eta in yeast. Only the C1 domain from Raf kinase enabled the chimeric enzyme to bind Rho1p when substituted for the native yeast domain. Accordingly, a chimeric Pkc1p carrying the C1 from Raf kinase, but not that from PKC eta, was able to partially complement the phenotypes of a yeast pkc1 deletion mutant. We interpret these data as further evidence that interaction with a small GTPase is the main regulatory function of the C1 domain in yeast.

Amino Acid Motifs↗
PubMed2001

Transient urethral obstruction predisposes to ascending pyelonephritis and tubulo-interstitial disease: studies in rats.

Chronic tubulo-interstitial disease, an important cause of end-stage renal disease, often results from the combined effects of a disturbed urinary outflow tract and urinary tract infection. Acute unilateral ureteral obstruction in rats rapidly induces foci of medullary necrosis, confined to the region of the papilla and fornices. This injury may provide a nidus for bacterial invasion and may invoke reactive and regenerative changes, ultimately leading to chronic pyelonephritis and tubulo-interstitial nephropathy. To explore this possibility, adult rats underwent renal morphological evaluation 2-7 days following transient 24-h unilateral ureteral obstruction. In some experiments the bladder was inoculated with bacteria (10(8)-10(9) cfu/ml Escherichia coli in 0.5 ml) after release of ureteral obstruction, with subsequent cultures obtained from the pelvis of both kidneys and from the urinary bladder. Morphologic evaluation of perfusion-fixed kidneys, 2-7 days after the release of 24-h ureteral obstruction disclosed papillary necrosis, urothelial proliferation, marked inner-stripe interstitial expansion, and fibrosis and proximal tubular (S3) dilatation. The lateral (perihilar region) was predominantly affected, with lesions spreading from the fornices. There was some progression of interstitial fibrosis during the postobstructive time course or following more prolonged ureteral obstruction. By contrast, infection hardly contributed to the tubulointerstitial changes. In rats subjected to infection, cultures were positive in all 15 postobstructive kidneys, as opposed to five contralateral kidneys (P < 0.0001). Viable counts from the postobstructive kidney were also higher than those from the contralateral side (79,000 +/- 12,000 vs 2900 +/- 1600 cfu/ml, mean +/- SEM, P < 0.0001), and were comparable to those obtained from the bladder (77,000 +/- 13,000 cfu/ml). We conclude that transient ureteral obstruction predisposes to ascending pyelonephritis and to tubulointerstitial disease. This vulnerability may relate to altered urodynamics and medullary tissue destruction.

Animals↗
PubMed2001

The yield of blood cultures in a department of emergency medicine.

This study sought to determine the yield of blood cultures drawn in the department of emergency medicine. The results of 730 blood cultures taken from 718 patients were retrospectively analysed. The total percentage of positive cultures was 9.7%. Only 3.4% of the blood cultures were classified as true bacteraemia and the rest as contaminants. The commonest type of isolate was coagulase-negative staphylococci (49%), which were considered contaminants in all cases. Other contaminants represented 13.2% of all the positive blood cultures. The following bacteria comprised the group of true bacteraemia: Escherichia coli (12.6%), Streptococcus pneumoniae (9.8%), viridans streptococci (7%), Staphylococcus aureus (2.8%), Bacteroides fragilis (2.8%), Moraxella species (1.4%) and Flavobacterium species (1.4%). Blood cultures were positive in 3.6% of patients with pneumonia and in 10% of patients with urinary tract infections. In patients with fever of unclear source blood cultures were positive in 3.1% of children between 0-36 months of age and in 1.1% of patients older than 16 years. As a whole, patients with positive blood cultures were clinically sicker, a higher percentage of them required admission to the hospital and had higher temperatures or rapidly fatal disease, compared with the group of patients with negative blood cultures. In order to improve the yield of blood cultures in febrile patients, first, better a priori identification of those subjects at high risk for bacteraemia will reduce the number of unnecessary blood cultures and second, sterile venipuncture techniques should be improved in order to reduce the number of contaminants.

Adolescent↗
PubMed2001

Acinetobacter baumannii at a tertiary-care teaching hospital in Jerusalem, Israel.

In a retrospective 10-year analysis of 3,536 patient-unique isolates, Acinetobacter baumannii imipenem susceptibility declined from 98.1 (1990) to 64.1% (2000), and ciprofloxacin susceptibility decreased from 50.5 to 13.1%. Imipenem median zone diameters decreased from 27. 7 (1997) to 18.8 mm (2000). No outbreaks were detected. Two clusters were identified for 41 strains genotyped by pulsed-field gel electrophoresis, but imipenem resistance was not clonal.

Acinetobacter↗
PubMed2001

Skin disease presenting as an outbreak of pseudobacteremia in a laboratory worker.

An outbreak of pseudobacteremia due to Streptococcus pyogenes (group A streptococci [GAS]) and methicillin-susceptible Staphylococcus aureus (MSSA) was traced to the venting procedure for aerobic bottles prior to their loading into the incubator of the BacT/Alert analyzer (Organon Teknika). Bacteria shed by a laboratory worker suffering from impetigo and cellulitis contaminated the aerobic bottles of 10 patients. All blood culture isolates, in addition to the isolates from the laboratory worker, were of the same GAS M and T types. All MSSA isolates from blood cultures and the index case's hands had the same lytic phage profile. Procedural breakdowns were identified in the laboratory. Bottles were vented outside the biological safety cabinet, gloves were not worn, and unprotected needles were used for the venting procedure. The source of the aspirated bacteria that contaminated the bottles was identified and the index case was treated promptly.

Bacteremia↗
PubMed2001

Multidrug-resistant Trichosporon asahii infection of nongranulocytopenic patients in three intensive care units.

Trichosporon asahii (Trichosporon beigelii) infections are rare but have been associated with a wide spectrum of clinical manifestations, ranging from superficial involvement in immunocompetent individuals to severe systemic disease in immunocompromised patients. We report on the recent recovery of T. asahii isolates with reduced susceptibility in vitro to amphotericin B (AMB), flucytosine, and azoles from six nongranulocytopenic patients who exhibited risk factors and who developed either superficial infections (four individuals) or invasive infections (two individuals) while in intensive care units. The latter two patients responded clinically and microbiologically to AMB treatment. All six isolates were closely related according to random amplified polymorphic DNA studies and showed 71% similarity by amplified fragment length polymorphism analysis, suggesting a common nosocomial origin. We also review the literature pertaining to T. asahii infections and discuss the salient characteristics of this fungus and recent taxonomic proposals for the genus.

Aged↗
PubMed2001

Enterobacter sakazakii infection in the newborn.

UNLABELLED: Enterobacter sakazakii, a Gram-negative bacillus, previously known as "yellow pigmented Enterobacter cloacae", is a rare cause of neonatal infection. We describe the detailed clinical presentation of two cases in whom E. sakazakii was isolated in our neonatal service during the course of 1 mo. These include one case of sepsis and meningitis complicated by cerebral infarction, and one case of sepsis. In addition, three cases of intestinal colonization were identified. The source of the organism was thoroughly sought and was found to be a blender in the milk kitchen that was used for preparation of the reconstituted powdered milk formula. CONCLUSION: Our paper adds clinical and laboratory information about the disease spectrum caused by this relatively rare organism and emphasizes the importance of a thorough search for the source of the infection.

Central Nervous System Infections↗
PubMed2000

Oncogenic insertional mutations in the P-loop of Ras are overactive in MAP kinase signaling.

Mutations of Ras with three extra amino acids inserted into the phosphate-binding (P) loop have been investigated both in vitro and in vivo. Such mutants have originally been detected as oncogenes both in the ras and the TC21 genes. Biochemical experiments reveal the molecular basis of their oncogenic potential: the mutants show a strongly attenuated binding affinity for nucleotides, most notably for GDP, leading to a preference for GTP binding. Furthermore, both the intrinsic as well as the GAP-stimulated GTP hydrolysis are drastically diminished. The binding interaction with GAP is reduced, whereas binding to the Ras-binding domain of the downstream effector c-Raf1 is not altered appreciably. Microinjection into PC12 cells shows the mutants to be as potent to induce neurite outgrowth as conventional oncogenic Ras mutants. Unexpectedly, their ability to stimulate the MAP kinase pathway as measured by a reporter gene assay in RK13 cells is much higher than that of the normal oncogenic mutant G12V. This characteristic was attributed to an increased stimulation of c-Raf1 kinase activity by the insertional Ras mutants.

Animals↗
PubMed2000

High rate of candidemia in patients sustaining injuries in a bomb blast at a marketplace: a possible environmental source.

In this study, a cluster of candidemia among patients sustaining injuries in a bomb blast at a marketplace was investigated by means of a multivariate analysis, a case-control study, and quantitative air sampling. Candidemia occurred in 7 (30%) of 21 patients (58% of those admitted to the intensive care unit [ICU]) between 4 and 16 days (mean, 12 days) after the injury and was the single most frequent cause of bloodstream infections. Inhalation injury was the strongest predictor for candidemia by multivariate analysis. Candidemia among the case patients occurred at a significantly higher rate than among comparable trauma patients injured in different urban settings, including a pedestrian mall (2 of 29; P=. 02), and among contemporary ICU control patients (1 of 40; P=.001). Air sampling revealed exclusive detection of Candida species and increased mold concentration in the market in comparison with the mall environment. These findings suggest a role for an exogenous, environmental source in the development of candidemia in some trauma patients.

Adolescent↗
PubMed2000

Phosphoinositide 3-kinase-dependent Ras activation by tauroursodesoxycholate in rat liver.

Ursodesoxycholic acid, widely used for the treatment of cholestatic liver disease, causes choleretic, anti-apoptotic and immunomodulatory effects. Here the effects on choleresis of its taurine conjugate tauroursodesoxycholate (TUDC), which is present in the enterohepatic circulation, were correlated with the activation of important elements of intracellular signal transduction in cultured rat hepatocytes and perfused rat liver. TUDC induced a time- and concentration-dependent activation of the small GTP-binding protein Ras and of phosphoinositide 3-kinase (PI 3-kinase) in cultured hepatocytes. Ras activation was dependent on PI 3-kinase activity, without the involvement of protein kinase C- and genistein-sensitive tyrosine kinases. Ras activation by TUDC was followed by an activation of the mitogen-activated protein kinases extracellular-signal-regulated kinase-1 (Erk-1) and Erk-2. In perfused rat liver, PI 3-kinase inhibitors largely abolished the stimulatory effect of TUDC on taurocholate excretion, suggesting an important role for a PI 3-kinase/Ras/Erk pathway in the choleretic effect of TUDC.

Animals↗
PubMed2000

Mitogenic signaling of Ras is regulated by differential interaction with Raf isozymes.

In the mitogenic signaling cascade interaction of Ras with Raf represents a critical step for the regulation of cell growth and differentiation. The major effector of Ras, the serine/threonine kinase Raf exists as three isoforms with different tissue distributions. We demonstrate that transient transfection of oncogenic Ha-Ras leads to a preferential activation of endogenous c-Raf-1 in HEK 293 cells as opposed to A-Raf. In vitro binding studies using purified Ras binding domains of Raf as well as in vivo bindings tests with full length molecules reveals significantly lower binding affinities of A-Raf to Ha-Ras as compared to other Raf isoforms. The Ras-binding interface of c-Raf differs from A-Raf by a conservative Arg to Lys exchange at residue 59 or 22 respectively. Mutational analysis reveals that this residue represents a point of isozyme discrimination: c-Raf-R59K binds Ha-Ras weaker than the wildtype, likewise A-Raf-K22R increases its affinity to Ha-Ras in vivo and in vitro. Differential binding affinities are reflected in downstream signaling. Immunecomplex kinase assays reveal that Ha-Ras mediated Raf activation is decreased for c-Raf-R59K and increased for A-Raf-K22R when compared to the respective wildtype forms. Thus our observations introduce a new level of isoform discrimination in Ras/Raf signaling as a functional consequence of a conservative amino acid exchange in the Ras binding domains.

Amino Acid Sequence↗
PubMed2000

Factors associated with nosocomial diarrhea and Clostridium difficile-associated disease on the adult wards of an urban tertiary care hospital.

A prospective survey of the adult inpatient population of an urban tertiary care hospital was conducted to determine factors associated with the development of nosocomial diarrhea and the acquisition of Clostridium difficile-associated disease. During the 3-month survey, 98 patients with nosocomial diarrhea were enrolled, and 38 controls were recruited. The controls were patients without diarrhea lying in beds adjacent to the affected patients. Factors significantly associated with nosocomial diarrhea were the administration of a special diet (P=0.02) and receipt of a greater number of different antibiotics (P=0.02). Among the 98 patients with diarrhea, Clostridium difficile toxin B was identified in the stool of 13. Factors found to be associated with the presence of toxin B as compared to other causes of nosocomial diarrhea were a greater number of individual antibiotics used during hospitalization (P=0.02) and receipt of a cephalosporin (P=0.03) or, more specifically, a third-generation cephalosporin (P=0.02). Among patients with nosocomial diarrhea, those who had toxin in their stool had a significantly higher total antibiotic burden (expressed as antibiotic days) than those with diarrhea due to other causes (P=0.01).

Adult↗
PubMed2000

Evaluation of chlorhexidine and povidone iodine activity against methicillin-resistant Staphylococcus aureus and vancomycin-resistant Enterococcus faecalis using a surface test.

Most published studies of the activity of biocides against methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant enterococci (VRE) have been based on suspension tests. This study was undertaken to provide information on the effect of chlorhexidine and povidone iodine on bacteria dried on to surfaces, a situation in which biocide activity is known to be reduced. The inactivation of MRSA (10 strains), methicillin-sensitive Staphylococcus aureus (MSSA, 10 strains), VRE (nine strains) and vancomycin-sensitive Enterococcus faecalis (VSE, 10 strains) by 0.5% aqueous chlorhexidine gluconate or 10% povidone iodine was evaluated by applying the European surface test method. Povidone iodine was equally active against resistant and sensitive strains of both species with microbicidal effects (ME), i.e. the log(10)concentration of micro-organisms compared with controls treated with distilled water, after 1.5 min of 3.14 and 3.49 for VRE and VSE respectively, and 3.47 and 3.78 for MRSA and MSSA. Chlorhexidine was equally active against VRE and VSE (ME 3.37 vs. 3. 56 after 7 min, respectively), but was significantly less active against MRSA as opposed to MSSA (ME 3.07 vs. 3.83 after 10 min, P= 0. 017).

Anti-Infective Agents, Local↗
PubMed1999

Thermodynamic and kinetic characterization of the interaction between the Ras binding domain of AF6 and members of the Ras subfamily.

Cellular signaling downstream of Ras is highly diversified and may involve many different effector molecules. A potential candidate is AF6 which was originally identified as a fusion to ALL-1 in acute myeloid leukemia. In the present work the interaction between Ras and AF6 is characterized and compared with other effectors. The binding characteristics are quite similar to Raf and RalGEF, i.e. nucleotide dissociation as well as GTPase-activating protein activity are inhibited, whereas the intrinsic GTPase activity of Ras is unperturbed by AF6 binding. Particularly, the dynamics of interaction are similar to Raf and RalGEF with a lifetime of the Ras. AF6 complex in the millisecond range. As probed by 31P NMR spectroscopy one of two major conformational states of Ras is stabilized by the interaction with AF6. Looking at the affinities of AF6 to a number of Ras mutants in the effector region, a specificity profile emerges distinct from that of other effector molecules. This finding may be useful in defining the biological function of AF6 by selectively switching off other pathways downstream of Ras using the appropriate effector mutant. Notably, among the Ras-related proteins AF6 binds most tightly to Rap1A which could imply a role of Rap1A in AF6 regulation.

Amino Acid Sequence↗
PubMed1999

Outbreak of bloodstream infections associated with dialysis machine waste ports in a hemodialysis facility.

Eight cases of gram-negative bloodstream infection without a clinically evident source occurred at a hemodialysis unit in Jerusalem between February and September 1997. All infections could be traced to three of the 13 dialysis machines in use. Epidemiological investigation, including pulsed-field gel electrophoretic characterization of organisms isolated from the patients and dialysis machines, implicated the Waste Handling Option system of the machines as the source of the infections. Discontinuation of the Waste Handling Option system use was associated with prompt cessation of the outbreak.

Adult↗
PubMed1999

Factors associated with pharyngeal carriage of Neisseria meningitidis among Israel Defense Force personnel at the end of their compulsory service.

In this 1 year cross-sectional study of personnel being discharged from compulsory military service, an available database of health-related information was used to examine the association of meningococcal carriage with socio-demographic factors. A representative, systematic sample of 1632 personnel was interviewed and had throat cultures taken. The overall meningococcal carriage rate was 16%. Serogroups B and Y accounted for 76% and 13% of the isolates respectively. In univariate analysis, carriage was associated with male gender (P < 0.0001), < 12 years school education (P = 0.002), smoking (P = 0.014), and service at a 'closed' base, reflecting greater interpersonal contact (P < 0.0001). In multivariate analysis, only service on a closed base and male gender retained significance. School education of < 12 years remained significant for females only. Variables not associated with carriage included number of siblings, intensity of smoking, and use of the contraceptive pill. In this setting, meningococcal carriage was associated with the type of base on which soldiers served; and smoking was not an independent risk factor for carriage.

Adult↗