Search results for "Polyacrylamide"
showing 10 items of 377 documents
Interaction ofEscherichia colihemolysin with biological membranes
2001
Escherichia coli hemolysin (HlyA) is a membrane-permeabilizing protein belonging to the family of RTX-toxins. Lytic activity depends on binding of Ca2(+) to the C-terminus of the molecule. The N-terminus of HlyA harbors hydrophobic sequences that are believed to constitute the membrane-inserting domain. In this study, 13 HlyA cysteine-replacement mutants were constructed and labeled with the polarity-sensitive fluorescent probe 6-bromoacetyl-2-dimethylaminonaphthalene (badan). The fluorescence emission of the label was examined in soluble and membrane-bound toxin. Binding effected a major blue shift in the emission of six residues within the N-terminal hydrophobic domain, indicating inserti…
Strong Cooperativity and Loose Geometry between CUB Domains Are the Basis for Procollagen C-Proteinase Enhancer Activity
2009
Procollagen C-proteinase enhancers (PCPE-1 and -2) specifically activate bone morphogenetic protein-1 (BMP-1) and other members of the tolloid proteinase family during C-terminal processing of fibrillar collagen precursors. PCPEs consist of two CUB domains (CUB1 and CUB2) and one NTR domain separated by one short and one long linker. It was previously shown that PCPEs can strongly interact with procollagen molecules, but the exact mechanism by which they enhance BMP-1 activity remains largely unknown. Here, we used a series of deletion mutants of PCPE-1 and two chimeric constructs with repetitions of the same CUB domain to study the role of each domain and linker. Out of all the forms teste…
Ultrastructure, fractionation and biochemical analysis of Cryptosporidium parvum sporozoites.
1999
Abstract Sporozoites of the apicomplexan parasite Cryptosporidium parvum were subjected to cell disruption and subcellular fractionation using a sucrose density step gradient. With this procedure, highly enriched preparations of the parasite membrane, the micronemes, dense granules and amylopectin granules were produced. No separate fraction containing rhoptries was obtained, however this organelle was found in defined fractions of the gradient, still associated with the apical tip of the sporozoites. Using negative staining, the internal structure of the micronemes was revealed by transmission electron microscopy. Micronemes and dense granules showed characteristic protein compositions by …
Deciliation: A stressful event for Paracentrotus lividus embryos.
1998
In this report, by using mono- and two-dimensional electrophoretic analysis, we demonstrate that deciliation on sea urchin embryos induces a stress response. Deciliation indeed causes not only the activation of ciliary subroutine, but also a transient decrease of bulk protein synthesis. This decrease is in agreement with our previous results on heat shock response in sea urchin, although deciliation does not induce the expression of the same main hsp set. We were able to characterize one main deciliation-stress protein of 40 kDa whose expression is transiently induced by deciliation and whose localisation is likely to be nuclear.
Ferric-reductase activities in Vibrio vulnificus biotypes 1 and 2.
1999
In this paper, the ferric-reductase activities of Vibrio vulnificus were investigated. This species comprises two biotypes pathogenic for humans and eels that are able to express different mechanisms for iron acquisition. All strains of both biotypes used in this study were able to reduce ferric citrate, irrespective of the iron levels in the growth medium. Some variation in the degree of reduction was observed among the strains, with the highest values corresponding to one acapsulated environmental strain of biotype 1. When cell fractions were tested, only those from periplasm and cytoplasm showed reductase activity whereas no activity was detected in membranes. Low temperatures inhibited …
Characterization of Different Deoxyribonucleases in Human Lymphocytes
1975
Abstract Deoxyribonucleases, Disc Electrophoresis, Lymphocytes Four groups of deoxyribonuclease activities from human lymphocytes have been characterized by deoxyribonuclease assay in DNA-containing polyacrylamide gels following their separation by disc-electrophoresis. All activities hydrolyse DNA endonucleolytically. One neutral deoxyribo nuclease found in the cytoplasmic fraction prefers native or UV-irradiated DNA over denatured DNA as substrate and is a 5′-monoester former. Two groups of acid deoxyribonuclease activities are detectable in the nuclear fraction. Both are 3′-monoester formers. One is as well active with denatured DNA as with native DNA, the other one shows the same activ…
Role of two operators in regulating the plasmid-borne raf operon of Escherichia coli
1994
The plasmid-borne raf operon encodes functions required for the inducible uptake and utilization of raffinose in Escherichia coli K12. The expression of three structural genes for alpha-galactosidase (rafA), Raf permease (rafB) and sucrose hydrolase (rafD) is negatively controlled by the binding of RafR repressor (rafR) to two operator sites, O1 and O2, that flank the -35 sequence of the raf promoter, PA. In vitro, O1 and O2 are occupied on increasing the concentration of RafR, without detectable preference for one site or the other or any indication of cooperative binding. Nucleotide substitutions at positions 3, 4 or 5 in an operator half-site prevented repressor binding, supporting a mod…
Plant progesterone 5β-reductase is not homologous to the animal enzyme. Molecular evolutionary characterization of P5βR from Digitalis purpurea
2007
Plants of the genus Digitalis produce cardiac glycosides, i.e. digoxin, which are widely used for congestive heart failure. Progesterone 5beta-reductase (P5betaR) is a key enzyme in the biosynthesis of these natural products. Here, we have carried out the purification and partial amino acid sequencing of the native P5betaR from foxglove (Digitalis purpurea), and isolated a cDNA encoding this enzyme. Similarly to other steroid 5beta-reductases, the recombinant P5betaR catalyzes the stereospecific reduction of the Delta(4)-double bond of several steroids with a 3-oxo,Delta(4,5) structure. The gene encoding P5betaR is expressed in all plant organs, and maximally transcribed in leaves and matur…
Characterization of a cDNA encoding RP43, a CUB-domain-containing protein from the tube of Riftia pachyptila (Vestimentifera), and distribution of it…
2000
A major 43kDa protein from the protective tube of Riftiapachyptila (Vestimentifera), named RP43, was partly microsequenced after isolation by SDS/PAGE from the protein fraction of tubes collected around the hydrothermal vents at the East Pacific Rise. On the basis of the partial peptide sequences obtained, experiments using reverse-transcriptase-mediated PCR and rapid amplification of cDNA ends led to the complete cDNA sequence. Analysis of deduced amino acid sequence of RP43 showed the presence of CUB domains (100–110-residue-spanning domains first reported in the complement subcomponents C1r/C1s, epidermal-growth-factor-related sea urchin protein and bone morphogenetic protein 1) that se…
Microdisc-electrophoretic study of deoxyribonucleases in cow snout epidermis
1983
Acid and neutral deoxyribonucleases (DNases) of the cow snout epidermis were investigated by the microdisc-electrophoresis of polyacrylamide gels containing highly polymerized DNA and by isoelectric focusing techniques. The nucleases were characterized with respect to their pH optimum. An acid DNase at pH 5.0 was detected as a single distinct band after the electrophoretic separation. After isoelectric focussing also, only one acid DNase activity with an isoelectric point (IP) of 6.2 was detectable. Neutral DNases at pH 7.4 were demonstrated as major and minor bands by their different electrophoretic mobilities. In the isoelectric focusing system also, two neutral DNases, a major one (IP, 4…