Biomedical subjects

R Alex

Publications and source records attributed to R Alex.

13 recordsLinked to original sources

PubMed1995

A basic tail increases repression by dimeric lac repressor.

Tetrameric Lac repressor achieves cooperative repression by binding simultaneously to O1 and to one of the auxiliary operators O2 or O3, thereby forcing the intervening DNA into a loop. Dimeric Lac repressor is not able to form DNA loops and consequently shows no cooperative repression. We constructed a dimeric Lac repressor mutant which exhibits increased repression to the lac operon that does not depend on specific operator-repressor-operator loops. This Lac repressor carries a synthetic tail of basic residues attached to its C terminus. With this construct, we observe an increase of the in vivo repression upon addition of auxiliary lac operators to a chromosomal lac operon controlled by O1. This suggests that the basic tail enables dimeric Lac repressor to enhance its repression by additional non-specific DNA contacts.

Amino Acid Sequence↗

PubMed1993

Single exchanges of amino acids in the basic region change the specificity of N-Myc.

We exchanged specific amino acids in the basic region of the murine N-Myc protein and tested the mutant proteins for their DNA binding specificity. The amino acids we exchanged were chosen in analogy to residues of the homologous basic regions of bHLH and bZIP proteins. Mutant N-Myc peptides were expressed in Escherichia coli and specific DNA binding was monitored by gel shift experiments. For this we used palindromic target sequences with systematic base pair exchanges. Several mutants with altered DNA binding specificity were identified. Amino acid exchanges of residues -14 or -10 of the basic region lead to specificity changes (we define leucine 402 of N-Myc as +1; comparable to GCN4 see (1)). The palindromic N-Myc recognition sequence 5'CACGTG is no longer recognized by the mutant proteins, but DNA fragments with symmetrical exchanges of the target sequence are. Exchanges at position -15 broaden the binding specificity. These data were used to build a computer based model of the putative interactions of the N-Myc basic DNA binding region with its target sequence.

Amino Acid Sequence↗

PubMed1992

Determination of the DNA sequence recognized by the bHLH-zip domain of the N-Myc protein.

The DNA-binding domain of the murine N-Myc protein, comprising the basic helix-loop-helix-zipper (bHLH-zip) region was expressed as a fusion protein in E. coli. The affinity purified glutathione-S-transferase-N-Myc fusion protein (GST-N-MYC) was used to select the N-Myc specific DNA-recognition motif from a pool of random-sequence oligonucleotides. After seven rounds of binding-site selection, specifically enriched oligonucleotides were cloned and sequenced. Of 31 individual oligonucleotides whose sequences were determined, 30 contained a common DNA-motif, defining the hexameric consensus sequence CACGTG. We confirm by mutational analysis that binding of the N-Myc derived bHLH-zip domain to this motif is sequence-specific.

Base Sequence↗

PubMed1991

[Biodegradable gentamicin-depot implants made of beta-tricalcium phosphate ceramics. 3. In vivo studies on drug release, tissue tolerance, and biodegradation].

In vivo drug release properties and biocompatibility of gentamicin-loaded controlled release implants made of beta-tricalcium phosphate ceramics designed for the local antibiotic treatment of bone infections were investigated. Controlled release pellets containing 0.4 and 0.8 mg of gentamicin were implanted into the femoral bone of rats. Drug release was measured from renal excretion over a time period of 3 weeks. The excretion pattern can be described by an initial phase of increased drug release was faster at higher drug loading. Drug release from glyceride-containing controlled release pellets occurs at a significantly slower rate than from drug-loaded pellets without glycerides. Histological studies after implantation of the pure ceramic pellets and the controlled release pellets into the bone tissue of rats and rabbits are showing a high tissue tolerance and the biodegradability of the implants. However, the glyceride-containing pellets are degraded at a slower rate than the pure ceramic pellets.

Animals↗

PubMed1988