Biomedical subjects

M T Nilsson

Publications and source records attributed to M T Nilsson.

4 recordsLinked to original sources

PubMed2000

Functional expression and affinity selection of single-chain cro by phage display: isolation of novel DNA-binding proteins.

A robust selection system affording phage display of the DNA-binding helix-turn-helix protein Cro is presented. The aim of the work was to construct an experimental system allowing for the construction and isolation of Cro-derived protein with new DNA-binding properties. A derivative of the phage lambda Cro repressor, scCro8, in which the protein subunits had been covalently connected via a peptide linker was expressed in fusion with the gene 3 protein of Escherichia coli filamentous phage. The phage-displayed single-chain Cro was shown to retain the DNA binding properties of its wild-type Cro counterpart regarding DNA sequence specificity and binding affinity. A kinetic analysis revealed the rate constant of dissociation of the single-chain Cro-phage/DNA complex to be indistinguishable from that of the free single-chain Cro. Affinity selection using a biotinylated DNA with a target consensus operator sequence allowed for a 3000-fold enrichment of phages displaying single-chain Cro over control phages. The selection was based on entrapment of phage/DNA complexes formed in solution on streptavidin-coated paramagnetic beads. The expression system was subsequently used to isolate variant scCro8 proteins, mutated in their DNA-binding residues, that specifically recognized new, unnatural target DNA ligands.

Bacteriophage lambda↗

PubMed1985

SEQ-ED: an interactive computer program for editing, analysis and storage of long DNA sequences.

The rapidly growing body of sequenced DNA demands efficient computer programs for its analysis and storage. The program described in this paper, SEQ-ED, has been designed to handle a large number of DNA sequences up to 200 kilobases [kb] long stored in a sequence library. In order to minimize the required storage space, the sequences are stored in a compressed format using three binary digits per base. In the development of this program, special care has been given to make it easy to use for molecular biologists without any previous computer experience.

Algorithms↗

PubMed1985

Purification of Epstein-Barr virus DNA polymerase from P3HR-1 cells.

The Epstein-Barr virus DNA polymerase was purified from extracts of P3HR-1 cells treated with n-butyrate for induction of the viral cycle. Sequential chromatography on DNA cellulose, phosphocellulose, and blue Sepharose yielded an enzyme preparation purified more than 1,300-fold. The purified enzyme was distinct from cellular enzymes but resembled the viral DNA polymerase in cells infected with herpes simplex virus type 1 or 2. The active enzyme had an apparent molecular weight of 185,000 as estimated by gel filtration on Sephacryl S-300. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a major polypeptide corresponding to a molecular weight of ca. 110,000. This polypeptide correlated with the catalytic function of the purified enzyme, whereas the other, less abundant polypeptides did not. By immunoblotting, the 110,000-molecular-weight polypeptide could be identified as a viral polypeptide. It could not be determined whether the native enzyme was composed of more than one polypeptide.

Animals↗

PubMed1983

Epstein-Barr virus and tumor promoters.

Epstein-Barr virus (EBV) can be maintained in lymphoblastoid cell lines. The infection is usually latent and the lack of a fully permissive cell have made the studies of the lytic functions of the virus difficult. In these cell lines virus production can be induced by addition of phorbol esters, n-butyrate, or nucleotide analogues. These compounds have been used to study EBV in several aspects: the latent and lytic functions of EBV. The possible use of this model system as an in vitro assay for tumor promoters. The mechanism behind the tumor promoter and n-butyrate induced activation of EBV.

B-Lymphocytes↗