Identification of the coding region for the putidaredoxin reductase gene from the plasmid of Pseudomonas putida

The first 12 NH₂-terminal amino acids of the Pseudomonas putida putidaredoxin reductase were shown to be Met-Asn-Ala-Asn-Asp-Asn-Val-Val-Ile-Val-Gly-Thr. Comparison of these data with the DNA sequence of the BamHI-HindIII 197-base fragment derived from the PstI 2.2-kb fragment obtained from the P. putida plasmid showed that the putidaredoxin reductase gene was downstream from the cytochrome P-450 gene and the intergenic region had the 24-nucleotide sequence TAAACACATGGGAGTGCGTGCTAA. The Shine-Dalgarno sequence GGAG was detected in this region. The initiating triplet for the reductase gene was GTG, which normally codes for valine, but in the initiating codon position codes for methionine. From the amino acid sequence and X-ray data comparisons with other flavoproteins, what appears to be the AMP binding region of the FAD can be recognized in the NH₂-terminal portion of the reductase involving residues 5–35.This article was presented during the proceedings of the International Conference on Macromolecular Structure and Function, held at the National Defence Medical College, Tokorozawa, Japan, December 1985.     

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) enhances antibody production and protein kinase activity in murine B cells

Treatment of murine spleen cells with 30 nM TCDD resulted in an approximately 3 fold increase in unstimulated antibody production after 3 days in culture. This response was not accompanied by increased cellular proliferation and may represent an effect of TCDD on B cell activation or differentiation. Since PMA is capable of activating B cells, presumably via PKC, we have compared the effects of PMA and TCDD on protein kinase activation and phosphorylation of endogenous proteins in a highly purified preparation of B cells. In contrast to a reduction of cytosolic PKC activity, the expected effect of PMA, TCDD caused an increase in basal kinase activity with no effect on PKC activity. Addition of either PMA or TCDD resulted in enhanced phosphorylation of a similar profile of proteins, including proteins of Mr 12.2, 14.6, 29.2, 52.3 and 62.7 KDa. Addition of TCDD also resulted in the increased phosphorylation of a protein of Mr 45.2, which was unaffected by PMA. Combined treatment with PMA and TCDD resulted in additive responses. The additive effects of PMA and TCDD suggest an interaction at the level of protein phosphorylation which is mediated by different kinases. Therefore, TCDD may be stimulating B cells via an early effect on an unidentified protein kinase.     

Evidence for lactate utilization for fetal lung glycogen synthesis

Fetal rabbit lungs from 23 day gestation animals were used to investigate the potential role of lactate as a substrate for fetal lung glycogen synthesis. Fetal lactate dehydrogenase activity was approximately twice that found in the adult lung, while the activity of phosphoenolpyruvate carboxykinase was elevated fourfold over the adult value. Pyruvate carboxylase activities were similar in both fetal and adult lungs. Studies employing fetal lung explants in organ culture indicated that the presence of both glucose and lactate may be necessary for glycogen accumulation in the developing fetal lung. These data support the hypothesis that lactate is an important precursor for fetal lung glycogen.     

Single channel kinetics of a glutamate receptor.

The glutamate receptor-channel of locust muscle membrane was studied using the patch-clamp technique. Muscles were pretreated with concanavalin A to block receptor-channel desensitization, thus facilitating analysis of receptor-channel gating kinetics. Single channel kinetics were analyzed to aid in identification of the molecular basis of channel gating. Channel dwell-time distributions and dwell-time autocorrelation functions were calculated from single channel data recorded in the precence of 10-4M glutamate. Analysis of the dwell time distributions in terms of mixtures of exponential functions revealed there to be at least three open states of the receptor-channel and at least four closed states. Autocorrelation function analysis showed there to be at least three pathways linking the open states with the closed. This results in a minimal scheme for gating of the glutamate receptor-channel, which is suggestive of allosteric models of receptor-channel gating.     

New and noteworthy South American species of Agalinis (Scrophulariaceae)

Agalinis bandeirensis is a Brazilian species distinguished by linear bracts, short pedicels, and villous stamens.Agalinis ramulifera, from southern Brazil, is distinguished by many, short, leafy branches and small flowers.Agalinis linarioides subsp.rojasi, from Paraguay, is distinguished by paniculate inflorescences, short calyx lobes, and broad corollas.Gerardia bangii, G. digitalis, andG. meyeniana are recognized as species ofAgalinis and appropriate transfers are made.     

A non-alphoid repetitive DNA sequence from human chromosome 21

Summary A non-alphoid repetitive DNA from human chromosome 22, consisting of a 48-bp motif, shows homology to both G-group chromosomes in the gorilla, thus indicating the presence of additional repeat family members on further human chromosomes. Therefore, we screened a chromosome-21-specific cosmid library using this repetitive sequence from chromosome 22 (D22Z3). Some 40–50 cosmid clones were positive in tests for hybridization. One of the clones giving the strongest signals was digested with EcoRI/PstI, which we knew to cut frequently within the repeats; this resulted in fragments containing repeat units only. The fragments were subcloned into plasmid vector pTZ 19. Sequence-analysis of a 500-bp insert showed ten copies of a 48-bp repeat similar to D22Z3, with about 15% sequence deviation from the chromosome 22 consensus sequence. In situ hybridization of the newly isolated recombinant established its chromosome 21 specifity at high stringency. Physical mapping by pulsed field gel electrophoresis placed this new repeat in close vicinity to the chromosome 21 alphoid repeat. No cross-hybridization with other mammalian genomes except for those of apes was observed. The locus has been designated D21Z2 by the Genome Data Base. A gel mobility shift assay indicated that this repetitive motif has protein-binding properties.     

Structure of an S layer on a pathogenic strain of Aeromonas hydrophila.

Negative staining revealed a tetragonal surface array (S layer) on all the members of a serogroup of Aeromonas hydrophila which possess high virulence for fish. The S layers were similar on all the strains examined, with unit cell dimensions of approximately 12 nm. A single representative strain, strain TF7, was selected for further analysis. Freeze-cleaved and etched preparations and sections for electron microscopy showed that the S layer was the outermost component of the cell envelope. This was confirmed by observation of thin sections. Computer-generated enhancements of the negatively stained micrographs showed the subunit organization to a resolution of less than 4 nm. Two structural units of identical lattice constants alternated in the array in both axes, and one of them was apparently dominant as the center of mass. The lesser unit was rotated 20 degrees from the dominant axes of symmetry and was formed by the junction of linker projections from a corner of the four components of the dominant unit. This interpretation was supported by finding that the array consists of a single polypeptide (molecular weight, 52,000). The unit cell as defined showed p4 symmetry, and a = b = 12.2 nm.     

New species of Aristolochia (Aristolochiaceae) from Peru

Aristolochia fosteri is a new species with unusual branched trichomes collected in Pasco near Oxapampa.Aristolochia hutchisonii is a prostrate species found in Amazonas and Cajamarca that is related toA. weberbaueri.Aristolochia barbouri is a narrow-leaved, herbaceous liana from Madre de Dios.     

Three new species of Elleanthus (Orchidaceae) from Central America

Elleanthus stolonifer andE. tillandsioides are new species in theE. poiformis complex of sectionChloidelyna.Elleanthus lentii is a new species in sectionStachydelyna. Elleanthus stolonifer is widespread in the mountains of Costa Rica and Panama;E. tillandsioides is found only in the lowland forests of southeastern Costa Rica;E. lentii is limited to the Cordillera de Guanacaste of Costa Rica.