Search results for "recombinant protein"
showing 10 items of 707 documents
Novel pathogenic mechanism of microbial metalloproteinases: liberation of membrane-anchored molecules in biologically active form exemplified by stud…
1996
Certain membrane-anchored proteins, including several cytokines and cytokine receptors, can be released into cell supernatants through the action of endogenous membrane-bound metalloproteinases. The shed molecules are then able to fulfill various biological functions; for example, soluble interleukin-6 receptor (sIL-6R) can bind to bystander cells, rendering these cells sensitive to the action of IL-6. Using IL-6R as a model substrate, we report that the metalloproteinase from Serratia marcescens mimics the action of the endogenous shedding proteinase. Treatment of human monocytes with the bacterial protease led to a rapid release of sIL-6R into the supernatant. This effect was inhibitable …
Crystallization and preliminary X-ray studies of mouse centrin1.
2005
The expression, purification, crystallization and preliminary X-ray diffraction studies of mouse centrin1 are reported. Centrins belong to a family of Ca{sup 2+}-binding EF-hand proteins that play a fundamental role in centrosome duplication and the function of cilia. To shed light on the structure–function relationship of these proteins, mouse centrin1 has been crystallized. The mouse centrin1 has been expressed in Escherichia coli as a GST-centrin fusion protein containing a thrombin protease cleavage site between the fusion partners. Two constructs with different linking-sequence lengths were expressed and purified. Thrombin cleavage yielded functional centrin1 and N-terminally extended …
Identification of N- and C-terminal Amino Acids of Lhca1 and Lhca4 Required for Formation of the Heterodimeric Peripheral Photosystem I Antenna LHCI-…
2002
Apoproteins of higher plant light-harvesting complexes (LHC) share considerable amino acid sequence identity/similarity. Despite this fact, they occur in different oligomeric states (i.e., monomeric, dimeric, and trimeric). As a step toward understanding the underlying structure requirements for different oligomerization behavior, we analyzed whether amino acids at the N- and C-termini of Lhca1 and Lhca4 are involved in the formation of the heterodimeric LHCI-730. Using altered proteins produced by deletion or site-directed mutagenesis for reconstitution, we were able to identify amino acids required for the assembly of LHCI-730. At the N-terminus of Lhca1, W4 is involved in heterodimerizat…
Mutation of a critical tryptophan to lysine in avidin or streptavidin may explain why sea urchin fibropellin adopts an avidin-like domain
1999
Sea urchin fibropellins are epidermal growth factor homologues that harbor a C-terminal domain, similar in sequence to hen egg-white avidin and bacterial streptavidin. The fibropellin sequence was used as a conceptual template for mutation of designated conserved tryptophan residues in the biotin-binding sites of the tetrameric proteins, avidin and streptavidin. Three different mutations of avidin, Trp-110-Lys, Trp-70-Arg and the double mutant, were expressed in a baculovirus-infected insect cell system. A mutant of streptavidin, Trp-120-Lys, was similarly expressed. The homologous tryptophan to lysine (W--K) mutations of avidin and streptavidin were both capable of binding biotin and bioti…
Recombinant NeutraLite Avidin: a non-glycosylated, acidic mutant of chicken avidin that exhibits high affinity for biotin and low non-specific bindin…
2000
AbstractA recombinant non-glycosylated and acidic form of avidin was designed and expressed in soluble form in baculovirus-infected insect cells. The mutations were based on the same principles that guided the design of the chemically and enzymatically modified avidin derivative, known as NeutraLite Avidin. In this novel recombinant avidin derivative, five out of the eight arginine residues were replaced with neutral amino acids, and two of the lysine residues were replaced by glutamic acid. In addition, the carbohydrate-bearing asparagine-17 residue was altered to an isoleucine, according to the known sequences of avidin-related genes. The resultant mutant protein, termed recombinant Neutr…
Recombinant avidin and avidin-fusion proteins.
2000
Both chicken egg-white avidin and its bacterial relative streptavidin are well known for their extraordinary high affinity with biotin (Kd approximately 10(-15) M). They are widely used as tools in a number of affinity-based separations, in diagnostic assays and in a variety of other applications. These methods have collectively become known as (strept)avidin-biotin technology. Biotin can easily and effectively be attached to different molecules, termed binders and probes, without destroying their biological activity. The exceptional stability of the avidin-biotin complex and the wide range of commercially available reagents explain the popularity of this system. In order by genetic enginee…
Molecular cloning and nucleotide sequence of chicken avidin-related genes 1-5.
1994
Using avidin cDNA as a hybridisation probe, we detected a gene family whose putative products are related to the chicken egg-white avidin. Two overlapping genomic clones were found to contain five genes (avidin-related genes 1–5, avrl-avr5), which have been cloned, characterized and sequenced. All of the genes have a four-exon structure with an overall identity with the avidin cDNA of 88–92%. The genes appear to have no pseudogenic features and, in fact, two of these genes have been shown to be transcribed. The putative proteins share a sequence identity of 68–78% with avidin. The amino acid residues responsible for the biotin-binding activity of avidin and the bacterial biotin-binding prot…
REGULATORY ELEMENTS OF THE LEUKAEMIA INHIBITORY FACTOR (LIF) PROMOTER IN MURINE BONE MARROW STROMAL CELLS
1999
Leukaemia inhibitory factor (LIF) plays an important role as a haematopoietically active cytokine. As described earlier in a murine model, interleukin 1 (IL-1) induced LIF mRNA and protein expression. We utilized the murine cell line +/+-1.LDA11 to further define regulatory mechanisms of LIF expression in bone marrow stromal cells. The production of LIF mRNA is stimulated by IL-1beta, TNF-alpha, and the cAMP analogue 8-bromoadenosine 3':5'-monophosphate (8BrcAMP). LIF mRNA expression is controlled at the transcriptional level. Different fragments from -542 to -45 bp 5' upstream of the transcriptional start site of the murine LIF gene were fused to the luciferase gene. All LIF-promoter lucif…
Interferon-alpha (IFN-alpha) inhibits granulocyte-macrophage colony-stimulating factor (GM-CSF) expression at the post-transcriptional level in murin…
1995
Recently it has been shown that IFN-alpha inhibits expression of GM-CSF in adherent cells of human long-term bone marrow cultures (LTBMC) stimulated with interleukin-1 (IL-1), tumour necrosis factor-alpha (TNF-alpha) or endotoxin. The murine bone marrow stromal cell line +/+(-1).LDA11 was used to further define regulatory mechanisms of IFN-alpha inhibition on GM-CSF expression. This cell line originated from a murine Dexter type culture and exhibits a preadipocytic phenotype. As in human LTBMC, we could demonstrate a inhibitory effect of IFN-alpha co-incubation on GM-CSF activity in serum-free supernatants of +/+(-1).LDA11 stromal cell cultures stimulated with IL-1 or TNF-alpha or the combi…
Easy removal of a large mucus plug with a flexible paediatric bronchoscope after administration of rhDNase (Pulmozyme).
2006
Mucoid impaction and plastic bronchitis are relatively rare disorders affecting patients of any age. Here we report the case of mucoid impaction developing in a four-year-old child as a complication of pneumonic infection treated four weeks prior. p.a. chest X-ray and fiberoptic bronchoscopy showed atelectasis and the location of a mucus cast in the left upper lobe. After extraction of the cast via the suction channel of the bronchoscope had failed, we administered 2.5 mg of undiluted recombinant human deoxyribonuclease (rhDNase) instilled directly over the affected area of occlusion for 15 minutes. This mucolytic agent mobilized the cast by reducing viscoelasticity and adherence of mucus. …