PubMed Health⌕ Search

Biomedical subjects

E Riise

Publications and source records attributed to E Riise.

16 recordsLinked to original sources
PubMed1999

Identification of patient-specific peptides for detection of M-proteins and myeloma cells.

We have taken advantage of the selection power of phage display technology to define specific peptide mimotopes that recognize individual M-proteins, isolated from patients with multiple myeloma. Preferred amino acid motifs of phages binding to M-proteins were identified in 6/9 patients investigated. Chemically synthesized peptides, corresponding to the phage-displayed peptide inserts, were used to verify the specificity of binding in competition assays. The peptides were able to bind to the M-proteins, as well as the myeloma cells, with high sensitivity and specificity. Employing simple immunological techniques, < 0.01 g/l of M-protein could be quantified, suggesting a novel way for monitoring minimal residual disease in the production of guidelines for adjusting or reintroducing conventional chemotherapy. The peptide mimotopes defined by this technology may be useful as tumour-specific targeting agents and as a tool for purging cells in autologous bone marrow transplantation.

Adult↗
PubMed1996

Inhibition of Serratia marcescens nuclease secretion by a truncated nuclease peptide.

The production of extracellular nuclease (Nuc) from the Serratia marcescens nucA chromosomal locus is inhibited in cells producing the N-terminal portion of Nuc from a multicopy plasmid. This inhibition in trans is not at the level of nucA expression, but rather at the level of secretion of the Nuc protein. Production of the periplasmic protein beta-lactamase (Bla) does not inhibit Nuc production unless fused to the nucA signal peptide and expressed from nucAp. Inhibition by either the truncated Nuc peptide (delta Nuc) or a Bla fusion protein is promoter specific and observed when expressed from nucAp; little inhibition is observed when the same protein is expressed from the lacZpo promoter-operator. This promoter specificity is also true for the secretion of Nuc itself.

Endodeoxyribonucleases↗
PubMed1993

Efficient method for constructing comprehensive murine Fab antibody libraries displayed on phage.

We have developed efficient methodologies for construction and expression of comprehensive phage display libraries of murine Fab antibody fragments in E. coli cells. Our methods optimize several critical steps of the polymerase chain reaction (PCR) amplification of transcripts of the re-arranged immunoglobulin genes and of their subsequent assembly and expression: Firstly, we have designed exhaustive sets of PCR primers of low degeneracy for the amplification of transcripts of the Fab region of the heavy and light-chain genes. These primers proved effective in amplification of Fab gene fragments from a large panel of hybridoma cell lines of different specificity and family sub-type. Secondly, we have developed a 'jumping PCR' technique that effectively assembled and recombined the amplified heavy and light-chain gene fragments into a bi-cistronic operon. Thirdly, we have constructed expression vectors for insertion of the combinatorial Fab gene-cassette in fusion with a truncated version of the phage surface protein, gIIIp. The heavy chain and the light chain-gIII fusion are transcribed as a polycistronic mRNA from the lacZ promoter and efficient transcriptional control is provided by wildtype lacI present on the vector. The utility of the system was demonstrated by isolating several antigen-binding clones from hybridomas and libraries made from immunized mice.

Animals↗
PubMed1990

Improved extraction method for avoiding the interference of ascorbic acid in the spectrophotometric determination of nitrite in meat products.

In minced meat products, ascorbic acid interferes in the usual spectrophotometric determination of nitrite by the diazotisation-coupling technique with sulphanilamide and N-(1-naphthyl)ethylenediamine. In a modified extraction method, the enzyme ascorbate oxidase was shown to diminish the interference of the acid. Within 10 min the enzyme was able to reduce the level of ascorbic acid in the minced meat specimens from 1000 p.p.m. down to the tolerance limit for interference. This appeared to be superior to the extraction method involving the use of water and was comparable to a previously described method based on the use of 0.01 M sodium hydroxide. As isoascorbic acid is not approved as a food additive in Norway no attempt was made to investigate the influence of this acid.

Ascorbate Oxidase↗
PubMed1990

Fermentation studies of the secretion of Serratia marcescens nuclease by Escherichia coli.

The secretion of a Serratia marcescens nuclease was followed by fermentation with Escherichia coli. A plasmid, p403-SD2, carrying a 1.3-kilobase-pair insert with a 0.4-kilobase-pair region upstream of the nuclease gene caused a growth-phase-regulated expression of nuclease in E. coli in the same way as that seen in S. marcescens. Deletion of the regulatory gene generating plasmid p403-Rsa1 resulted in a constitutive expression of the nuclease. Anaerobiosis stimulated the expression from p403-SD2 in stationary growth phase by a factor of 10 compared with expression stimulated by cultivation in aerobic conditions; no such effect was found for plasmid p403-Rsa1. Different nutritional factors caused the expression level and the amount of extracellular nuclease to vary more when nuclease was expressed from plasmid p403-SD2 than when it was expressed from plasmid p403-Rsa1. A correlation between the regulatory gene and the extracellular secretion of nuclease is proposed.

Bacteriological Techniques↗
PubMed1989

Purification and characterization of a Serratia marcescens nuclease produced by Escherichia coli.

The primary structure and physical chemical properties were determined of a nuclease expressed and secreted by Escherichia coli. The plasmid p403-SD2 carried a DNA sequence isolated from Serratia marcescens encoding the enzyme. During cultivation of the E. coli cells, 85% of the enzyme was released to the growth medium. The enzyme was purified and exhibited a single band with a molecular weight about 30,600 daltons on SDS-PAGE similar to nuclease isolated from S. marcescens. The amino acid composition and the amino acid sequence determined directly confirmed the primary structure of 245 amino acids predicted from the DNA sequence, and, in addition, the two disulfide bridges were assigned. Several physical chemical properties were examined. The ability of the enzyme to cross the outer membrane is proposed to depend upon the formation of the proper structures during the folding process.

Amino Acid Sequence↗
PubMed1987

Two-level factorial screening for influence of temperature, pH, and aeration on production of Serratia marcescens nuclease.

Two high-nuclease-yielding mutants of Serratia marcescens, derived by chemical mutagenesis (W280, W355), and two strains with the pBR322 plasmid 403-SD2, carrying a nuclease gene and a chloramphenicol resistance gene [Escherichia coli CSH50(403-SD2) and S. marcescens CH30(403-SD2)] were investigated for nuclease production in a factorial shake flask experiment, with temperature (30 and 37 degrees C), pH (with or without CaCO3 tablets), and aeration (with or without baffles) as variable conditions. Yields varied 10-fold depending on the conditions investigated.

Aerobiosis↗
PubMed1986

Purification and characterization of the CopB replication control protein, and precise mapping of its target site in the R1 plasmid.

The CopB regulatory loop from plasmid R1 has been analyzed. The CopB protein was partially purified, but proteolytic activity in vitro resulted in the recovery of two molecular forms of the polypeptide. Both of these acted as repressors of the repA promoter and had identical activities. The smaller of the proteins was found to be the result of a specific cleavage in the normal in vivo translation product. The active form of the CopB protein is most likely a tetramer, which binds to a DNA region overlapping the repA promoter that also contains a stretch of dyad symmetry. Footprinting analysis and mutant analysis (including nucleotide sequence determination) identified this binding site within 20-25 base pairs. In agreement with in vivo results the binding between CopB and its target site is moderate compared with other operons like lac and trp.

Base Sequence↗
PubMed1985

Transcription and its regulation in the basic replicon region of plasmid R1.

The transcriptional units in the basic replicon of plasmid R1 were defined by means of gene fusions. It was found that in the wild-type plasmid there is one large mRNA encoding both the control factor copB and the positive replication factor repA. A second, internal transcription initiation site, the repA promoter, is usually repressed by the copB protein, and is therefore only of significance in the absence of this control factor. By induction of the repA promoter through gradual dilution of the copB repressor it was shown that translation of repA-mRNA, controlled by the copA-RNA, is significantly increased only when the rate of repA transcription is above a certain level. No indication was found for a possible interference from convergent copA transcription on repA transcription.

Base Sequence↗
PubMed1982

Molecular cloning and functional characterization of a copy number control gene (copB) of plasmid R1.

Deletions or insertions in the copB gene of plasmid R1 result in a copy mutant phenotype. The wild-type copB gene has been cloned on various plasmid vectors. The presence of such chimeric plasmids reduced the copy number of R1 copB mutant plasmids to normal or subnormal levels, indicating the expression of a trans-acting inhibitor activity from the copB chimeras. However, the cloned copB gene did not affect the copy number of wild-type R1, and no incompatibility was exerted by the cloned copB gene against wild-type R1 (or R100). Although the copB gene is not normally required for the incompatibility exerted by copA, it is shown that the CopB function is required for expression of incompatibility by the copA gene from some types of chimeric plasmids. Mutant plasmids that have lost both Cop functions replicate in an uncontrolled fashion.

Cloning, Molecular↗
PubMed1981

The genes for the eight subunits of the membrane bound ATP synthase of Escherichia coli.

The genes for the eight subunits of the membrane bound ATP synthase of Escherichia coli (Ca++, Mg++ dependent ATPase, EC 3.6.1.3) were mapped through genetic, physical and functional analysis of specialized transducing phages lambda asn (von Meyenburg et al. 1978). The ATP synthase genes, designated atp1, are located at 83.2 min in a segment of the chromosome between 3.5 and 11.3 kb left (counterclockwise) of the origin of replication oriC. The counterclockwise order of the genes for the eight subunits, the expression of which starts from a control region at 3.5 kb-L, was found to be: a, (c, b, delta), alpha, gamma, (epsilon, beta) which in the notation of Downie el al. (1981) reads atp B (EFH) A G (C D). The analysis was in part based on the isolation of new types of atp (unc, Suc-) mutations. We made use of the fact that specialized transducing phages lambda asn carrying oriC can establish themselves as minichromosomes rendering asnA cells Asn+, and that the resulting Asn+ cells grow slowly if the lambda asn carries part or all of the atp operon. Selecting for fast growing strains mutations were isolated on the lambda asn which either eliminated atp genes or affected their expression ("promoter" mutations). The relationship between these atp mutations and the cop mutations of Ogura et al. (1980), which also appear to map in front of or within the atp genes, is discussed.

Bacteriophage lambda↗
PubMed1981

Isolation and characterization of lambda phages carrying the basic replicon of the resistance plasmid R1.

Specialized transducing lambda phages, lambda oriR1, harboring DNA from the resistance plasmid R1drd-19 and its copy mutant pKN103 were isolated. From measurements of CCC-DNA content it is concluded that upon infection the phages can establish themselves as self-replicating plasmids in recA hosts lysogenic for lambda. It is thought that this bypassing of lambda immunity is due to the presence of the R1 origin of replication. The plasmids are sensitive to the incompatibility expressed by plasmid R1. This has been shown mainly by transduction of lambda oriR1 into recipients containing R1 plasmids or plasmid pBR322 carrying the basic replicon. We were able to demonstrate that a copy mutant of plasmid R1 was insensitive to copA+, but sensitive to the concerted action of PstI fragments F1 and F2. This mutant was previously assumed to be of dominant type. Physical mapping of the lambda oriR1 derivatives verified that they carry the basic replicon of plasmid R1. The plasmids are not stably maintained, but are lost in a frequency of 1%-2% per cell generation, which is consistent with their lack of the R1par region.

Bacteriophage lambda↗
PubMed1978

Origin of replication, oriC, or the Escherichia coli chromosome on specialized transducing phages lambda asn.

Specialized transducing phages lambda asn harboring chromosomal DNA and genetic markers on either side of the asn gene were isolated. Phages carrying chromosomal DNA counterclockwise of the asn gene can upon infection establish themselves as self-replicating plasmids in asn, recA hosts lysogenic for lambda. It is concluded that this bypassing of normal lambda immunity is due to the presence of the chromosomal replication origin, oriC, in this class of phages. Genetic analysis and the determination of restriction endonuclease cleavage patterns of the different lambda asn lead to the allocation of oriC within 1.5 megadaltons of the asn gene towards the uncA, uncB genes at 82 min on the genetic map of E. coli. The clockwise order of genes on the chromosomes is found to be: bglB, (pst, glmS), (uncA, uncB), oriC, asn, trkD, rbs, rrnC, ilv.

Chromosome Mapping↗