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Biomedical subjects

M Rider

Publications and source records attributed to M Rider.

14 recordsLinked to original sources
PubMed2000

Regulation of protein kinase D by multisite phosphorylation. Identification of phosphorylation sites by mass spectrometry and characterization by site-directed mutagenesis.

Activation of the serine/threonine kinase, protein kinase D (PKD/PKC mu) via a phorbol ester/PKC-dependent pathway involves phosphorylation events. The present study identifies five in vivo phosphorylation sites by mass spectrometry, and the role of four of them was investigated by site-directed mutagenesis. Four sites are autophosphorylation sites, the first of which (Ser(916)) is located in the C terminus; its phosphorylation modifies the conformation of the kinase and influences duration of kinase activation but is not required for phorbol ester-mediated activation of PKD. The second autophosphorylation site (Ser(203)) lies in that region of the regulatory domain, which in PKC mu interacts with 14-3-3tau. The last two autophosphorylation sites (Ser(744) and Ser(748)) are located in the activation loop but are only phosphorylated in the isolated PKD-catalytic domain and not in the full-length PKD; they may affect enzyme catalysis but are not involved in the activation of wild-type PKD by phorbol ester. We also present evidence for proteolytic activation of PKD. The fifth site (Ser(255)) is transphosphorylated downstream of a PKC-dependent pathway after in vivo stimulation with phorbol ester. In vivo phorbol ester stimulation of an S255E mutant no longer requires PKC-mediated events. In conclusion, our results show that PKD is a multisite phosphorylated enzyme and suggest that its phosphorylation may be an intricate process that regulates its biological functions in very distinct ways.

Alanine↗
PubMed2000

Clinical evaluation of a novel chromogenic agar dipslide for diagnosis of urinary tract infections.

The performance of a novel dipslide (DipStreak; Novamed, Israel) consisting of chromogenic agar (Uriselect 3; Sanofi Pasteur, France) and blood agar media was evaluated prospectively and compared to that of conventional urine culture for the diagnosis of urinary tract infection. A total of 1070 clean-catch urine specimens obtained from 251 hospitalized patients and 819 outpatients were processed. The overall performance of the DipStreak was as follows: sensitivity, 95.7%; specificity, 99.2%; agreement, 89.8%; accuracy, 98%; positive predictive value, 98.5%; and negative predictive value, 97.7%. A total of 270 urine specimens were positive by both DipStreak and conventional culture. The chromogenic agar allowed rapid identification of organisms in 211 (78.1%) cultures, while isolates in the other 59 (21.9%) cultures remained unidentified. The results indicate that the DipStreak device coupled with the Uriselect 3 agar represents a convenient and accurate method for inoculation of urine specimens, quantitation of bacteria, diagnosis of significant bacteriuria, and presumptive identification of isolates.

Bacteria↗
PubMed1996

Iron repressible outer membrane proteins of Moraxella bovis and demonstration of siderophore-like activity.

Moraxella bovis (strain Epp 63), grown in RPMI 1640 medium supplemented with desferrioxamine mesylate (0.05 mg/ml) resulted in cell free culture supernatants with an increased chromeazurol-S response indicating the presence of high affinity iron binding ligand(s). Supernatants of cultures where growth occurred in tryptic soy broth, RPMI 1640, or RPMI 1640-desferrioxamine supplemented with ferrous sulfate (10 micrograms/ml) were negative on the chromeazurol-S test. Growth of M. bovis in RPMI 1640 or RPMI 1640-desferrioxamine medium induced the expression of previously unrecognized outer membrane proteins whose expression was repressed when the medium was supplemented with iron and which were not produced when growth occurred in tryptic soy broth.

Bacterial Outer Membrane Proteins↗
PubMed1995

Production of recombinant human brain type I inositol-1,4,5-trisphosphate 5-phosphatase in Escherichia coli. Lack of phosphorylation by protein kinase C.

The dephosphorylation of inositol 1,4,5-trisphosphate (InsP3) to inositol 1,4-bisphosphate is catalyzed by InsP3 5-phosphatase. The coding region of human brain type I InsP3 5-phosphatase was expressed as a fusion protein with the maltose-binding protein (MBP) in Escherichia coli, using the pMAL-cR1 vector. The relative molecular mass of the purified fusion protein (MBP-InsP3-5-phosphatase) was approximately M(r) 85,000 as analysed by SDS/PAGE. The yield was about 10 mg fusion protein/l lysate. After cleavage from MBP with factor Xa, the specific activity of recombinant 5-phosphatase was 120-250 mumol.mg-1.min-1. The molecular mass of purified protein by SDS/PAGE was M(r) 43,000. The activity was inactivated by p-hydroxymercuribenzoate. The possibility that protein kinase C might phosphorylate InsP3 5-phosphatase was tested on the purified 43,000 M(r) protein. In this study, we show that recombinant 5-phosphatase is not a substrate of protein kinase C.

Animals↗
PubMed1994

Partial characterization of Streptococcus suis type 2 hemolysin.

Streptococcus suis type 2 was evaluated for hemolysin production. Supernatants of S. suis type 2 grown in Todd-Hewitt broth were assayed for hemolytic activity by a photometric assay. Twenty-two additional serotypes of S. suis (1,3 to 22, and 1/2) were evaluated for hemolysin production; nine of them (1/2, 1, 4, 5, 14, 15, 17, 19, and 20) were positive. The effects of temperature, atmosphere, centrifugation, sonication, chemicals, bovine serum albumin, fetal calf serum, and enzymes on S. suis type 2 hemolysin activity were studied. Maximum hemolysis occurred after incubation in RPMI 1640 medium at 40 degrees C in 6% CO2 and after growth in Todd-Hewitt broth at 37 degrees C under anaerobic conditions. Hemolytic activity was absent after the addition of fetal calf serum and decreased after the addition of trypsin or amylase. However, treatment of erythrocytes with amylase or trypsin prior to incubation with supernatant also resulted in a decrease in hemolytic activity. The addition of bovine serum albumin caused increased hemolytic activity. Dipyridyl and EDTA had negligible effects on hemolysis. Hemolytic S. suis type 2 culture supernatant injected intraperitoneally failed to cause death in BALB/c mice. Data from our study indicate that S. suis type 2 hemolysin is a secreted or loosely cell bound, thermolabile molecule whose activity is growth condition dependent.

Animals↗
PubMed1991

Anti-idiotypic antibodies mimic bovine viral diarrhea virus antigen.

Polyclonal rabbit anti-idiotypic antibodies (anti-ids) against two neutralizing murine monoclonal antibodies (mAbs) specific to a bovine viral diarrhea virus (BVDV) glycoprotein, 53 kDa, were produced, purified, and characterized. Each anti-id inhibited the binding of its respective mAb to BVDV antigen in a competitive ELISA and blocked the immunoprecipitation of the 53 kDa protein by the mAb. The anti-ids also inhibited the virus-neutralizing activity of their homologous mAbs. These results suggest that the anti-ids bear an internal image of a BVDV antigen and mimic neutralizing epitopes on the 53 kDa protein. Treatment of MDBK cells with the anti-ids inhibited BVDV infection, indicating that they block a cellular component, such as a virus receptor, required for virus adsorption or entry. Inhibition of the homologous mAb and lack of inhibition of the heterologous mAb indicate that the anti-ids are specific for the unique antigen-binding sites on the mAbs.

Animals↗
PubMed1991

Healthy cities: the Indiana model of community development in public health.

While the Institute of Medicine's Report on The Future of Public Health provokes debate on public health issues in the United States, the Healthy Cities movement is one approach to addressing many of these issues. Healthy Cities Indiana began in 1988 and adapts the European and Canadian healthy cities experience within the sociopolitical context of Indiana and the United States. Six Indiana cities are collaborating with Indiana University School of Nursing and the Indiana Public Health Association in a process of urban health promotion. The overall healthy cities process is presented including: city commitment, formation of healthy city committee, community leadership development, city action, provision of data-based information to policy makers, and action research and evaluation. Each step in the healthy cities process is described, highlighting examples from the Indiana experience, including an analysis of selected data describing the cities. The facilitators of a healthy city provide support to city leaders in other cities interested in becoming involved in the healthy cities process. Implications of healthy cities for health educators and other health professionals are suggested.

Community Participation↗
PubMed1988

Rod/cone dysplasia in Irish setters. Presence of an altered rhodopsin.

On the basis of the amino acid sequence of bovine rhodopsin, a series of peptides from the C-terminus (Rhod-4 and Rhod-1) and external loops (Rhod-10) were synthesized. Rabbit antisera to these peptides recognize the rhodopsin molecule in whole retina from 8-week-old normal and affected rcdl (rod/cone-dysplasic) Irish setters (8- and 4-weeks-old). When the rhodopsin content was equalized by using a solid-phase radioimmunoassay, the reaction with anti-peptide antisera to the C-terminal octapeptide (residues 341-348) is severely decreased in the rcdl-dog retinas. The results of mixing experiments suggest that this is not due to proteolytic clipping of the rhodopsin C-terminus from the affected dogs. Treatment of retinas with 1.0 mM-NaF, a phosphatase inhibitor, or pretreatment with alkaline and acid phosphatases does alter the reaction of the rhodopsin with anti-rhodopsin antisera. This suggests that the decreased reaction of the affected rhodopsin with the anti-peptide antisera may partially result from differences in intrinsic rhodopsin phosphorylation. However, since the reaction of rcdl retinas cannot be restored to that of the normals, these results suggest that the rhodopsin molecule from the rcdl dogs may be structurally altered in other ways.

Alkaline Phosphatase↗
PubMed1988

Characterization of nifH mutations of Klebsiella pneumoniae.

Nucleotide changes in the nifH gene of Klebsiella pneumoniae were identified by DNA cloning and sequencing of six selected mutant strains. The strains were UN60, C-640-GC----TGC; UN116, C-67-TC----TTC; UN117, G-688-AG----AAG; UN1041, CG-302-C----CAC; UN1678, GC-713-C----GTC; and UN1795, G-439-AG----AAG. Their corresponding amino acid substitutions were UN60, Arg-214----Cys; UN116, Leu-23----Phe; UN117, Glu-230----Lys; UN1041, Arg-101----His; UN1678, Ala-238----Val; and UN1795, Glu-147----Lys. Results from Western and Northern blots of the mutant strains showed significant reductions in both steady-state levels of the accumulated Fe protein and nifH mRNA during derepression in the presence of serine. The relative specific activities of the nitrogenases in strains UN60, UN1041, and UN1795 were less than 2% of the wild type, whereas those in UN116, UN117, and UN1678 were between 28 and 40% of the wild type during enhanced derepression with serine. The residues of Arg-101 (UN1041), Glu-147 (UN1795), and Arg-214 (UN60) were invariant in sequences of a dozen diazotrophs that have been examined thus far. In UN1041, in which Arg-101 of the Fe protein was replaced by His, the Fe protein had a larger apparent molecular weight than that of the other strains on sodium dodecyl sulfate-gel electrophoresis, as detected with rabbit antiserum raised against the C-terminal peptide of the wild-type Fe protein. The reduced levels of nifH mRNA in point mutant strains suggests that nifH (the gene or gene product) may be involved in self-regulation. mRNA transcripts of different sizes were detected when a nifH-specific probe, CCKp2003, was used in the Northern blot hybridization.

DNA, Bacterial↗
PubMed1977

Investigation of adhesive properties of dental composite materials using an improved tensile test procedure and scanning electron microscopy.

A standardized tension test was used to evaluate the adhesive properties of several composite materials when used on both dentin and enamel specimens. The nature of the test surfaces was examined by roughness tests using a Talysurf machine and also in more detail by means of a scanning electron microscope. Poor results were obtained for the adhesion of composite materials to dentin whereas good retention to enamel was obtained.

Adhesiveness↗