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K Hofmann

Publications and source records attributed to K Hofmann.

At least 91 records · Page 5Linked to original sources
PubMed1996

Performance and application potential of ion-molecule reaction mass spectrometry (IMR-MS) in the analysis of complex gas mixtures.

The mild ionisation technique of IMR mass spectrometry in combination with a tuneable cell for the charge transfer reaction allows to study mixtures of small molecules with low fragmentation and chemical selectivity. Temporal resolution down to 100 ms and quasisimultaneous multicomponent trend analyses can be combined in a flexible way responding to changing analytical requirements. The research instrument of ATOMIKA provides digital control over a range of parameters and ionisation sources. Its performance is characterised in several applications ranging from purity tests over the detection of small oxygenate organic molecules to more complex reaction systems of heterogeneous catalytic performance testing.

Journal Article↗
PubMed1996

A flexible motif search technique based on generalized profiles.

A flexible motif search technique is presented which has two major components: (1) a generalized profile syntax serving as a motif definition language; and (2) a motif search method specifically adapted to the problem of finding multiple instances of a motif in the same sequence. The new profile structure, which is the core of the generalized profile syntax, combines the functions of a variety of motif descriptors implemented in other methods, including regular expression-like patterns, weight matrices, previously used profiles, and certain types of hidden Markov models (HMMs). The relationship between generalized profiles and other biomolecular motif descriptors is analyzed in detail, with special attention to HMMs. Generalized profiles are shown to be equivalent to a particular class of HMMs, and conversion procedures in both directions are given. The conversion procedures provide an interpretation for local alignment in the framework of stochastic models, allowing for clear, simple significance tests. A mathematical statement of the motif search problem defines the new method exactly without linking it to a specific algorithmic solution. Part of the definition includes a new definition of disjointness of alignments.

Algorithms↗
PubMed1996

A consensus motif in the RFX DNA binding domain and binding domain mutants with altered specificity.

The RFX DNA binding domain is a novel motif that has been conserved in a growing number of dimeric DNA-binding proteins, having diverse regulatory functions, in eukaryotic organisms ranging from yeasts to humans. To characterize this novel motif, we have performed a detailed dissection of the site-specific DNA binding activity of RFX1, a prototypical member of the RFX family. First, we have performed a site selection procedure to define the consensus binding site of RFX1. Second, we have developed a new mutagenesis-selection procedure to derive a precise consensus motif, and to test the accuracy of a secondary structure prediction, for the RFX domain. Third, a modification of this procedure has allowed us to isolate altered-specificity RFX1 mutants. These results should facilitate the identification both of additional candidate genes controlled by RFX1 and of new members of the RFX family. Moreover, the altered-specificity RFX1 mutants represent valuable tools that will permit the function of RFX1 to be analyzed in vivo without interference from the ubiquitously expressed endogenous protein. Finally, the simplicity, efficiency, and versatility of the selection procedure we have developed make it of general value for the determination of consensus motifs, and for the isolation of mutants exhibiting altered functional properties, for large protein domains involved in protein-DNA as well as protein-protein interactions.

Amino Acid Sequence↗
PubMed1996

Methodological and clinical evaluation of two automated enzymatic immunoassays as compared with a radioimmunoassay for neuron-specific enolase.

We evaluated the clinical and methodological features of the neuron-specific enolase radioimmunoassay (NSE RIA) (Pharmacia = Ph) with the neuron-specific enolase enzyme immunoassay (NSE EIA) on the ES 700 (Boehringer Mannheim = BM) and the NSE EIA on the Cobas Core System (Roche = Ro). A total of 253 serum samples obtained from 37 healthy persons, 45 patients with benign lung diseases, 124 patients with lung cancer (42 with small cell lung cancer, 23 with adenocarcinoma, 21 with squamous cell carcinoma, 11 with large cell carcinoma, and 27 with unknown histology), 34 with lung metastases, 7 patients with sarcoma and 6 patients with malign lymphatic diseases were stored at -80 degrees C and assayed retrospectively. The intra- and inter-assay imprecisions were lower for the automatized test systems than for the RIA. Correlation between the EIA's and the RIA was better for NSE (Ro) than for NSE (BM) (BM/Ph: r = 0.93 and slope = 0.54; Ro/Ph: r = 0.95, slope = 0.79), but weaker than the correlation between the two EIA's: over the whole range r = 0.96, neuron-specific enolase < 50 micrograms/l: r = 0.97, neuron-specific enolase < 20 micrograms/l: r = 0.92. Fixing the specificity at 95% versus benign lung diseases we found a cut off value of 11.9 micrograms/l for NSE RIA (Ph), 15.9 micrograms/l for NSE EIA (BM) and 13.5 micrograms/l for NSE EIA (Ro). Based on this specificity of 95% versus benign lung diseases as the clinically relevant reference group, the sensitivity for NSE RIA was 32% for all lung cancer and 45% for small cell lung cancer, for NSE EIA (BM) 35% for all lung cancer and 43% for small cell lung cancer, the NSE EIA (Ro) had a sensitivity of 42% for all lung cancer and 57% for small cell lung cancer. In a follow-up study of two patients with small cell lung cancer a good comparability for all three assays in the kinetics, but a marked difference in the neuron-specific enolase value levels was found. The results show that the NSE EIA (Ro) on Cobas Core system is the most sensitive assay for the detection of small cell lung cancer.

Enzyme-Linked Immunosorbent Assay↗
PubMed1996

A sequence similarity search algorithm based on a probabilistic interpretation of an alignment scoring system.

We present a probabilistic interpretation of local sequence alignment methods where the alignment scoring system (ASS) plays the role of a stochastic process defining a probability distribution over all sequence pairs. An explicit algorithms is given to compute the probability of two sequences given and ASS. Based on this definition, a modified version of the Smith-Waterman local similarity search algorithm has been devised, which assesses sequence relationships by log likelihood ratios. When tested on classical examples such as globins or G-protein-coupled receptors, the new method proved to be up to an order of magnitude more sensitive than the native Smith-Waterman algorithm.

Algorithms↗
PubMed1995

Mammalian homologues of Caenorhabditis elegans unc-13 gene define novel family of C2-domain proteins.

The unc-13 gene in Caenorhabditis elegans is essential for normal presynaptic function and encodes a large protein with C1- and C2-domains. In protein kinase C and synaptotagmin, C1- and/or C2-domains are regulatory domains for Ca2+, phospholipids, and diacylglycerol, suggesting a role for unc-13 in regulating neurotransmitter release. To determine if a similar protein is a component of the presynaptic machinery for neurotransmitter release in vertebrates, we studied unc-13 homologues in rat. Molecular cloning revealed that three homologues of unc-13 called Munc13-1, -13-2, and -13-3 are expressed in rat brain. Munc13s are large, brain-specific proteins with divergent N termini but conserved C termini containing C1- and C2-domains. Specific antibodies demonstrated that Munc13-1 is a peripheral membrane protein that is enriched in synaptosomes and localized to plasma membranes but absent from synaptic vesicles. Our data suggest that the function of unc-13 in C. elegans is conserved in mammals and that Munc13s act as plasma membrane proteins in nerve terminals. The presence of C1- and C2-domains in these proteins and the phenotype of the C. elegans mutants raise the possibility that Munc13s may have an essential signaling role during neurotransmitter release.

Amino Acid Sequence↗
PubMed1995

The death domain motif found in Fas (Apo-1) and TNF receptor is present in proteins involved in apoptosis and axonal guidance.

The interaction of Fas (Apo-1) and TNF receptor-1 with their respective ligands can lead to cell death. The so-called death domain, a sequence motif present in the cytoplasmic portion of the two receptors, has been identified as a critical structural element involved in signal transduction that leads to apoptosis. Here we describe several additional proteins which contain a death domain. Novel members of this family include proteins known to be implicated not only in apoptosis but also in neuron guidance.

Amino Acid Sequence↗
PubMed1995

The S. pombe cdc15 gene is a key element in the reorganization of F-actin at mitosis.

The S. pombe cdc15 gene is essential for cell division. cdc15ts mutants do not form a septum, but growth and nuclear division continue, leading to formation of multinucleate cells. The earliest step in septum formation and cytokinesis, rearrangement of actin to the center of the cell, is associated with appearance of hypophosphorylated cdc15p and formation of a cdc15p ring, which colocalizes with actin. Loss of cdc15p function impairs formation of the actin ring. The abundance of cdc15 mRNA varies through the cell division cycle, peaking in early mitosis before septation. Expression of cdc15 in G2-arrested cells induces actin rearrangement to the center of the cell. These data implicate cdc15p as a key element in mediating the cytoskeletal rearrangements required for cytokinesis.

Actins↗
PubMed1995

The rsp5-domain is shared by proteins of diverse functions.

A novel, unusually small, and highly conserved domain of modular intracellular proteins is described. The domain was first recognized as three repeats in the yeast rsp5 gene product and named thereafter. The rsp5 protein is thought to interact with nuclear proteins but also contains a C2 domain typical for cytoplasmic proteins. Further analyses revealed several additional occurrences of this domain in diverse protein classes, including cytoplasmic signal transduction proteins, gene products interacting with the transcription machinery, structural proteins like dystrophin, and a putative RNA helicase.

Amino Acid Sequence↗
PubMed1995

Changes in histological structure and physiochemical properties of raw, cured, smoked pork loins resulting from freezing near cryoscopic temperature.

The aim of the study was to determine the effects of unintended freezing during cold storage (deep chilling conditions) and low freezing temperatures (-10 degrees C) of raw pork loins, cured and smoked, on their histological structure and some physicochemical characteristics. The experimental products after freezing at -10 degrees C, were stored at the same temperature for 0, 2, 4 and 6 days, thawed thereafter and immediately deep chilled at near cryoscopic temperature (-3 degrees C) for 0, 2, 4 or 6 days. These conditions were changing the histological structures of pork loin, the water holding capacity (WHC) was reduced by frozen storage but improved by deep chilling; the maximum shear force and energy were increased. The more the freezing at -10 degrees C was prolonged, the more unfavourable were the changes in structure, WHC and tenderness. Deep chilling at -3 degrees C after thawing of the experimental material on the other hand favourably affected the parameters measured. Freezing at -10 degrees C and deep chilling at -3 degrees C did not affect the electrophoretic picture of pork loin proteins nor the water activity. Freezing itself had no influence on WHC. Changes were due to storage.

Animals↗
PubMed1995

Escherichia coli K12 relA strains as safe hosts for expression of recombinant DNA.

Most Escherichia coli K12 strains survive for a relatively long time outside the laboratory. Under the same conditions the isoallelic E. coli K12 relA mutants die faster because they lack the stringent response. The killing rate is increased by using a plasmid-encoded suicide system consisting of the phage T7 lysozyme gene driven by the E. coli alkaline phosphatase gene promoter (phoA). Cells containing this system were rapidly and effectively killed as soon as phosphate was made limiting. The combination of the chromosomal relA mutation and a conditional suicide system of this type provides an effective means of biological containment for recombinant E. coli strains.

Alkaline Phosphatase↗
PubMed1994

Human neutral amino acid transporter ASCT1: structure of the gene (SLC1A4) and localization to chromosome 2p13-p15.

Screening for cDNAs encoding proteins similar to the sodium-coupled glutamate transporter GLAST1 led to the isolation of a cDNA clone coding for a protein that turned out to be identical to the recently described neutral amino acid transporter ASCT1. The new member of the GLAST-related transporter family does not transport glutamate or aspartate but alanine, serine, cysteine, and threonine instead. The expressed sequence tag EST02446, a short cDNA sequence found in the course of a large-scale sequencing project of human brain-derived cDNAs, showed significant similarity to the eukaryotic glutamate transporter GLAST1 and was therefore used as probe in the search for further glutamate transporter cDNAs. Fragments of the cDNA were used for the isolation and characterization of human ASCT1 genomic clones. The ORF of 1572 bp encoding 524 amino acid residues is distributed over 8 exons, which span at least 40 kb of human chromosomal DNA. The ASCT1 gene locus was assigned to chromosome 2p13-p15 by chromosomal in situ suppression (CISS) studies. The gene structure is not related to any other previously characterized transporter gene. In contrast to the genes of the sodium-coupled nonglutamate neurotransmitter transporters, it shows no obvious correspondence between intron/exon structure and transmembrane organization. The transcription start site in human liver tissue was determined by primer extension analysis to be located 291 bp upstream of the initiating ATG codon. The DNA region immediately upstream of the transcription start lacks any TATA or CAAT boxes but contains several binding sites for the transcription factors Sp1 and Egr-1.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1994

Efficient use of lactose for the lac promoter-controlled overexpression of the main antigenic protein of the foot and mouth disease virus in Escherichia coli under fed-batch fermentation conditions.

Derivatives of the lac promoter (tac, pac, rac) belong to the strongest bacterial promoters which are frequently used for the induced overexpression of foreign genes in Escherichia coli. However, their use in fermentation processes is strongly restricted because of the high cost of the inducer iso-propyl-beta-D-thiogalactopyranoside (IPTG). The aim of this work was to investigate the possibility of using lac-derived promoters in high cell density processes resulting in a high yield of the induced recombinant protein if glucose is the main carbon and energy source. Lactose is tested as inducer of the main antigenic coat protein (VP1) of the foot and mouth disease (FMD) virus in a T7-RNA polymerase expression system. It was shown that lactose is able to induce the expression of the recombinant gene to an amount of the VP1 protein corresponding to 20% of the total cell protein.

Antigens, Viral↗
PubMed1993

Molecular cloning and gene organization of the mouse mitochondrial 3,2-trans-enoyl-CoA isomerase.

3,2-trans-enoyl-CoA isomerase (mECI, E.C. 5.3.3.8) is the key enzyme of mitochondrial beta-oxidation of unsaturated fatty acids. A mouse cDNA clone spanning the entire coding region of mECI was isolated and sequenced. Subsequently, two overlapping genomic clones containing the complete mECI gene were isolated and characterized. The mouse mECI cDNA comprises an open reading frame of 867 bp, encoding a protein of 32 kDa. The mECI gene, spanning about 15 kb, consists of seven exons. Multiple transcription starts were determined by primer extension experiments. Knowledge of the gene organization and availability of genomic clones for mouse mECI will facilitate the study of unsaturated fatty acid metabolism in normal and pathological states.

Amino Acid Sequence↗