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K Neugebauer

Publications and source records attributed to K Neugebauer.

6 recordsLinked to original sources
PubMed1985

Differential responses of L5 and rat primary muscle cells to factors in rat brain extract.

Crude brain extract (100,000 g supernate from newborn or fetal rat brain homogenate) was studied for its effects on the number and distribution of acetylcholine receptors (AChRs) on myotubes of the L5 cloned myogenic cell line and compared to that of rat primary cultures. Gamma counting, light autoradiography and scanning electron microscopic autoradiography were used. We found that the L5 cells responded to the brain extract with an increase in the average AChR site density (2-5-fold) and with an increase in AChR clustering. Clustering was manifested by both an increase in the number of AChR clusters and in the ratio of receptor site density within clusters relative to that between clusters. The increase in average AChR site density was shown to be due to an increase in the rate of AChR insertion into the surface membrane with little change in the rate of receptor degradation. As also previously reported, the rat myotubes had a similar clustering response but only a very slight (approximately 1.2-fold) increase in average AChR site density. The surface area of myotubes was also increased slightly (approximately 1.2-1.3-fold) by the brain extract. Autoradiography viewed by scanning EM was found to be very useful in illustrating the shape and distribution of the receptor clusters. After the brain extract was fractionated on Sephadex G-200, the fractions with greatest clustering activity could be separated from those causing predominantly an increase in receptor site density. Increased receptor site density was primarily produced by the low molecular weight fractions (less than 12 kD), whereas the strongest (but not exclusive) effect on clustering was produced by the high molecular weight fractions (greater than 140 kD). Furthermore, the two cell types assayed had different sensitivity to the different factors. L5 cells responded to both the high and low molecular weight factors while rat primary cells are sensitive primarily to the high molecular weight factors.

Animals↗
PubMed1981

Penicillinase from Bacillus licheniformis: nucleotide sequence of the gene and implications for the biosynthesis of a secretory protein in a Gram-positive bacterium.

The gene for the penicillinase from B. licheniformis has been cloned in a functional stat on a 1.5 kb DNA fragment and its nucleotide sequence has been determined. A sequence of 307 amino acid residues is infered for the penicillinase precursor. Of these 34 amino acids precede the sequence of the secreted form of the enzyme. This peptide extension shows the features of a signal for secretion and also provides the hydrophobic anchor for the membrane-bound form of the enzyme.

Bacillus↗
PubMed1981

Bacillus licheniformis penicillinase synthesized in Escherichia coli contains covalently linked fatty acid and glyceride.

DNA sequence analysis of the structural gene for Bacillus licheniformis penicillinase has revealed a tetrapeptide sequence of Leu-Ala-Gly-Cys within the NH2-terminal part of the precursor form of penicillinase (penicillin amido-beta-lactamhydrolase, EC 3.5.2.6). The same tetrapeptide occurs in the signal sequence of the prolipoprotein of Escherichia coli, and the cysteine residue in the tetrapeptide of prolipoprotein is modified to form glyceride-cysteine which becomes the NH2 terminus of Braun's lipoprotein. On the basis of labeling, with [2-3H]glycerol, [3H]palmitate, [35S]methionine, and [35S]sulfuric acid, of an E. coli strain lysogenic for a lambda vector containing the penicillinase gene from B. licheniformis and of immunoprecipitation with rabbit antisera against purified B. licheniformis penicillinase, we conclude that B. licheniformis penicillinase synthesized in E. coli contains covalently linked glyceride and fatty acid. These results strongly suggest the operation of a modification system in E. coli, and presumably in other Gram-negative bacteria, which results in the formation of a glyceride-cysteine residue if the proper peptide sequence is present in the signal sequence of membrane proteins.

Bacillus↗
PubMed1980

Conversion of bacteriophage fd into an efficient single-stranded DNA vector system.

Single-stranded DNA vectors were constructed in vitro by insertion of various DNA fragments into the Intergenic Region of the single-stranded DNA phage fd. These inserts introduce into the phage genome unique cleavage sites for restriction nucleases which are suited for sticky joining in cloning experiments. Since these sites are usually located within genes coding for antibiotic resistance, inactivation of a resistance gene by insertion can be used as a marker for the successful cloning of a DNA fragment. Resistance genes also allow to select for recombinant DNA phages and to minimize the loss of DNA inserts which otherwise becomes significant above an insert size of about one kb. Cloning of several DNA fragments is described and strand separation of double-stranded DNA fragments by means of cloning into fd DNA is given as an example for application of single-stranded DNA vectors.

Bacteriophages↗
PubMed1978

Transposition of a DNA sequence determining kanamycin resistance into the single-stranded genome of bacteriophage fd.

Derivatives of bacteriophages fd which transduce kanamycin resistance were selected after growth of the phage in an E. coli strain that carried transpoon 5 (Tn5). Different clones of transducing phage and their DNAs were characterized by gel electrophoresis, electron microscopy, and by their ability to multiply in the absence of helper phage. Integration of the intact transposon into the full size phage genome was correlated with an increase in size of the phage particle from 0.95 mu to 1.7 mu, and with the appearance in the phage DNA of the stem loop structure characteristic for single-stranded Tn5 DNA. In non-defective phages the site of insertion was mapped by heteroduplex analysis within the intergenic region of the phage genome. Defective transducing phages were characterized as an insertion of Tn5 into a phage gene, and/or as a partial deletion or duplication of phage and transposon DNA. The size of the transducing phage from different defective clones varied from 0.6 mu to 3.0 mu and was directly proportional to the DNA content. These results demonstrate that filamentous bacteriophage are highly capable to replicate and package very different amounts of foreign DNA.

Coliphages↗