The nomenclature of the Pacific herring, Clupea pallasü Valenciennes, 1847

Confusion over the nomenclature of the Pacific herring is shown to have stemmed from the comparison of it with White Sea herring which was mistakenly assumed to be conspecific with Atlantic herring.     

Formaldehyde-derived tetrahydroisoquinolines and tetrahydro-β-carbolines in human urine

Human urine samples were examined for the occurrence of formaldehyde-derived tetrahydroisoquinolines and tetrahydro-β-carbolines generated by condensation of the methanol oxidation product with biogenic amines. Positive results were obtained for the tryptamine condensation product 1,2,3,4-tetrahydro-β-carboline and the serotonine condensation product 6-hydroxy-1,2,3,4-tetrahydro-β-carboline as well as for the condensation products with tyramine, dopamine, adrenaline and noradrenaline 1,2,3,4-tetrahydroisoquinoline, 6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline, N-methyl-4,6,7-trihydroxy-1,2,3,4-tetrahydroisoquinoline, 4,6,7-trihydroxy-1,2,3,4-tetrahydroisoquinoline, and the metabolite 6-methoxy-7-hydroxy-1,2,3,4-tetrahydroisoquinoline. Negative results were obtained for N-methyl-1,2,3,4-tetrahydroisoquinoline and 6,7-di-methoxy-1,2,3,4-tetrahydroisoquinoline, N-methyl-1,2,3,4-tetrahydro-β-carboline, 6-methyl-1,2,3,4-tetrahydro-β-carboline, and 6-methoxy-1,2,3,4-tetrahydro-β-carboline in samples of chronic alcoholics as well as in the urine of healthy volunteers. No correlation between alcohol ingestion or state of alcoholization could be demonstrated.     

An approach to integrated antibody production: Coupling of fluidized bed cultivation and fluidized bed adsorption

Continuous culture may be an efficient way of producing proteins which are susceptible to secondary processing in the course of a fermentation process. Short residence times in these systems support the production of correctly assembled proteins by avoiding substrate limitations and product inhibitions and also minimize the contact of sensitive bioproducts with degrading enzymes. Thus products of increased stability and integrity are obtained from continuous processes. The downstream process following continuous culture has to be adapted to the specific conditions of continuous fermentations, e.g. large liquid volumes and diluted process solutions. In this paper an approach is shown how a fluidized bed adsorption as first recovery operation may be coupled directly to a continuous production. Immobilized hybridoma cells are cultivated in porous glass microcarriers in a continuous fluidized bed process, the cell containing harvest is purified by fluidized bed adsorption using an agarose based cation exchange matrix. By this coupled mode of operation the large biomass containing harvest volume resulting from the continuous cultivation may be applied directly to a fluidized chromatographic matrix without prior clarification, leading to a particle free and initially purified product solution of reduced volume. In an experimental setup a bench-scale fluidized bed bioreactor of 25 ml carrier volume was coupled to a fluidized bed adsorption column operated with 300 ml of adsorbent. This configuration yielded up to 20 mg of monoclonal antibody per day in a cell free solution at fourfold concentration and fivefold purification. The process was run for more than three weeks with consistent product output.The help of H. Schmitz, A. Bader, J. Gätgens and M. Halfar during the experiments is gratefully acknowledged. This work was partially funded by the ministry of science and research of the Federal Republic of Germany within the project Stoffumwandlung mit Biokatalysatoren.     

Analysis of the protein kinase activity of moss phytochrome expressed in fibroblast cell culture

In the moss Ceratodon purpureus a phytochrome gene encodes a phytochrome type (PhyCer) which has a C-terminal domain homologous to the catalytic domain of eukaryotic protein kinases (PKs). PhyCer exhibits sequence conservation to serine/ threonine as well to tyrosine kinases. Since PhyCer is expressed very weakly in moss cells, to investigate the proposed PK activity of PhyCer, we overexpressed PhyCer transiently in fibroblast cells. For this purpose we made a chimeric receptor, EC-R, which consists of the extracellular, the membrane-spanning and the juxtamembrane domains of the human epidermal growth-factor receptor (EGF-R) linked to the PK catalytic domain of PhyCer (CerKin). The expression of EC-R in transiently transfected cells was confirmed with antibodies directed against the extracellular domain of EGF-R or against CerKin. Both EGF-R and EC-R were immunoprecipitated from lysates of overexpressing cells with antibodies against the extracellular domain of EGF-R. Phosphorylation experiments were performed with the immunoprecipitates and the phosphorylation products were subjected to phosphoamino acid analysis. Phosphorylation products specifically obtained with EC-R-transfected cells exhibit phosphorylation on serine and threonine residues. In EC-R transfected cells the endogenous EGF-R showed enhanced phosphorylation of serine and threonine residues compared to EGF-R immuno-precipitated from control cells. Although CerKin is closest to the catalytic domain of a protein tyrosine kinase from Dictyostelium discoideum, EC-R does not appear to phosphorylate tyrosine residues in vitro. From our data we conclude that PhyCer carries an active PK domain capable of phosphorylating serine and threonine residues.Abbreviations CerKin protein kinase catalytic domain of PhyCer - EC-R chimeric receptor consisting of the extracellular, the membrane spanning and the juxtamembrane domains of the human epidermal growth factor receptor (EGF-R) linked to the protein kinase catalytic domain of PhyCer - EGF-R epidermal growth factor receptor - mAb monoclonal antibody - PhyCer phytochrome gene in Ceratodon encoding a phytochrome type which has a C-terminal domain homologous to the catalytic domain of eucaryotic protein kinases - PK protein kinase - PVDF polyvinyl difluoride - Ser serine - Thr threonine - Tyr tyrosine Dr. Patricia Algarra was supported by the Alexander von Humboldt Foundation, Germany. This work was supported by the Deutsche Forschungsgemeinschaft (DFG), Bonn, Germany.     

Analysis of human V H gene repertoire expression in peripheral CD19+ B cells

Using CD19 B-cell selection and polymerase chain reaction-amplified cDNA libraries, we analyzed the peripheral immunoglobulin heavy chain variable repertoire of three healthy adult donors. Here we report that most of the CD19+ circulating B cells expressed unmutated V _H-D-J_H rearrangements. By specific V _H family hybridization, we show that V _H gene family utilization in the periphery roughly corresponds to the complexity of these families in the germline and appears to be relatively constant among the analyzed subjects. However, sequence data of clones picked at random from one IgM cDNA library reveals that in spite of this random utilization, the V _H gene expression in naive circulating B cells is highly biased towards the expression of a limited set of V _H genes. As previously reported by others, this restricted mechanism is also found for the D and J _H segments.The nucleotide sequence data reported in this paper have been submitted to the GenBank/EMBL nucleotide sequence database and have been assigned the accession numbers Z47213-Z47243 and Z47349     

Differential accumulation of the transcripts of 22 novel protein kinase genes in Arabidopsis thaliana

22 novel members of the Arabidopsis thaliana protein kinase family (AKs) were identified by using degenerate oligonucleotide primers directed to highly conserved amino acid sequences of the protein kinase (PK) catalytic domain. Of these 22 genes, 16 turned out to carry intron sequences. Homologies of AK sequences were detected to S-locus receptor protein kinases (SRKs) from Brassica spp., to SRK-like PKs from maize and A. thaliana and to several other receptor PKs from A. thaliana. Sequence similarity was also detected to Ca²⁺-dependent PKs (CDPKs) from rape and soybean, to SNF1 and to CDC2 homologues. The genomic organization and the accumulation of the mRNAs from these 22 AK genes were investigated.     

Purification of monoclonal antibodies from whole hybridoma fermentation broth by fluidized bed adsorption

To achive the coarse purification of a monoclonal antibody from whole hybridoma fermentation broth a fluidized bed cation exchange process was used. The procedure consisted of application of the crude sample and washing of the bed in a fluidized mode and elution in a fixed bed mode. A completely clarified eluate was obtained with purification factors between 4 and 8 and a concentration of the desired product (monoclonal antibody) by a factor of more than 3 was achived. Thus, a combination of the three early steps of the downstream process clarification, concentration and coarse purification was possible. Two different materials were tested: a commercially available agarose-based matrix (Stream-line-SP), and a self-derivatized material based on controlled-pore glass (Bioran). Initial experiments were performed to describe the fluidization of the glass material. Comparison with the agarose material showed several differences, the agarose matrix allowing liquid flow closer to plug flow than the glass material. Increased backmixing in the liquid phase was detected when fluidizing the glass adsorbent compared with the agarose-based matrix. Despite this fact, comparison of the two materials with respect to antibody binding and elution demonstrated a similar performance. (c) 1995 John Wiley & Sons, Inc.     

Reproduction and continuity of chloroplasts in spermatophytes

Without the continuity of chloroplasts there would be no higher life. Chloroplast reproduction and continuity can be studied in the guard cells of stomata, utilizing the result of reproduction, viz., the number of chloroplasts. An annotated list of such numbers from 261 families of spermatophytes is provided. For each family and many subfamilies or tribes are given the numbers of genera and species investigated as well as the ranges and medians of 1909 generic and 6161 specific chloroplast numbers, predominantly from my own countings. The driving forces behind the reproduction and continuity of chloroplasts are reviewed and discussed in the light of some figures of the list and other evidence. The results support the hypothesis that nuclear DNA replication paves the way for chloroplast reproduction and that the nuclear DNA amount is instrumental in ensuring the continuity of chloroplasts throughout meristematic cell successions.