Immobilized cells: behaviour of carrageenan entrapped yeast during continuous ethanol fermentation

Summary Data of cell concentration, viability and microscopic observation of cell distribution inside carrageenan immobilized yeast beads are reported. Results were obtained from a continuous packed-bed reactor performing alcoholic fermentation and the main observations made on cell activity are in agreement with the fermentation profiles inside the fermenter.     

Discrimination of alfalfa weevil strains by allozyme analysis

Allozyme profiles of eastern weevils (Beltsville, Maryland; Washington Co., Illinois), western weevils (Logan and St. George, Utah), and Egyptian weevils (Yuma, Arizona; Westmorland, California) were compiled by acrylamide gel electrophoresis. Twenty-two gene loci from 12 enzymes (ACPH, ADH, AMY, AO, EST, GOT, G-6PDH, MDH, ME, SOD, TYR, XDH) were analyzed. Mean heterozygosity of these populations was 0.231, with an average proportion of polymorphic loci of 0.536. The mean genetic distance of all weevil populations was 0.033 and the fixation index was 0.024. Diagnostic loci were found which could distinguish western weevils from eastern and Egyptian weevils. The small genetic distance between the eastern and Egyptian weevils suggests that they may be the same strain and are certainly different from the western weevil strain. Based on this and other evidence, we conclude that all weevil strains in the United States are Hypera postica (Gyllenhal), and that the use of H. brunneipennis (Boheman) for the Egyptian alfalfa weevil of North America should be discontinued.

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Résumé Les allozymes d'Hypera de l'est des USA (Beltsville, Maryland; Washington Co., Illinois), de l'ouest des USA (Logan et St Georges, Utah) et égyptiens (Yuma, Arizona; Westmorland, Californie) ont été analysés par électrophorèse sur gel d'acrylamide. L'étude a porté sur 22 loci de 12 enzymes (ACPH, ADH, AMY, AO, EST, GOT, G-6PDH, MDH, ME, SOD, TYR, XDH). L'hétérozygotie moyenne de la population était 0.231, avec une moyenne de loci polymorphes de 0.536. La distance génétique moyenne de l'ensemble des populations était de 0.033 et l'indice de fixation de 0.024. Des loci caractéristiques ont été trouvés qui pourraient permettre de distinguer les Hypera occidentaux des orientaux et des égyptiens. La faible distance génétique entre les Hypera orientaux et égyptiens suggère qu'ils appartiennent à la même souche et sont certainement différents des occidentaux. A partir de cela et d'autres éléments, nous concluons que tous les Hypera des USA sont H. postica Gyllenhal et que l'utilisation d'H. brunneipennis Boheman pour désigner les Hypera égyptiens d'Amérique du Nord doit être abandonnée.

     

Streptomycin-resistance of Euglena gracilis chloroplasts: identification of a point mutation in the 16S rRNA gene in an invariant position.

We sequenced the chloroplast 16S rRNA gene of two Euglena gracilis mutants which contain streptomycin-resistant chloroplasts (Smr 139.12/4 and Smr 139.20/2). These mutants are known to contain a single intact rrn operon per circular chloroplast genome. Nucleotide sequence comparison between a 16S rRNA gene of wild type Euglena gracilis, strain Z, with streptomycin-sensitive chloroplasts, and the 16S rRNA gene of both Smr-strains reveals a single base change (C to T) at position 876. This position is equivalent to the invariant position 912 of the E. coli 16S rRNA gene. The analogous position is also conserved in all chloroplast small subunit RNA genes from lower and higher plants sequenced so far. Light dependent protein synthesis with purified chloroplasts from streptomycin-resistant cells is not inhibited by streptomycin. Based on the results reported here we postulate linkage between the observed point mutation on the 16S rRNA gene and streptomycin-resistance of chloroplast 70S ribosomes.     

Nucleotide sequence of a 'truncated rRNA operon' of the Euglena gracilis chloroplast genome.

An extra 16S rRNA gene (s-16S rDNA) from the Euglena gracilis chloroplast genome and several hundred positions of its flanking regions have been sequenced. The structural part has 1486 positions and is to 98% homologous in its sequence with the 16S rRNA gene in functional chloroplast rRNA operons. Sequences of about 200 positions upstream and 15 positions downstream of the structural part of the s-16S rRNA gene region are highly homologous with corresponding parts in the functional operon. Neither tRNA genes (A1a, I1e) nor parts of the 23S and 5S rRNA genes are found within 557 positions after the 3' end of the s-16S rRNA gene, i.e., the 330 bp homology, observed in electron microscopic studies of heteroduplexes (4), between the s-16S rDNA downstream region and the 6.2 kb repeated segment containing the functional rRNA operon, must be due to a DNA stretch in the interoperon spacer. A structural model of the "truncated rRNA operon" is presented. Results from S-1 endonuclease analysis suggest that the s-16S rDNA region is probably not transcribed into stable s-16S rRNA.     

Renal Cortical Basolateral Na+/HCO− 3 Cotransporter: IV Characterization and Localization with Polyclonal Antibodies

We have previously partially purified the basolateral Na⁺/HCO⁻ ₃ cotransporter from rabbit renal cortex and this resulted in a 400-fold purification, and an SDS-PAGE analysis showed an enhancement of a protein band with a MW of approximately 56 kDa. We developed polyclonal antibodies against the Na⁺/HCO⁻ ₃ cotransporter by immunizing Dutch-belted rabbits with a partially purified protein fraction enriched in cotransporter activity. Western blot analysis of renal cortical basolateral membranes and of solubilized basolateral membrane proteins showed that the antibodies recognized a protein with a MW of approximately 56 kDa. The specificity of the purified antibodies against the Na⁺/HCO⁻ ₃ cotransporter was tested by immunoprecipitation. Solubilized basolateral membrane proteins enriched in Na⁺/HCO⁻ ₃ cotransporter activity were incubated with the purified antibody or with the preimmune IgG and then reconstituted in proteoliposomes. The purified antibody fraction caused a concentration-dependent inhibition of the Na⁺/HCO⁻ ₃ cotransporter activity, while the preimmune IgG failed to elicit any change. The inhibitory effect of the antibody was of the same magnitude whether it was added prior to (inside) or after (outside) reconstitution in proteoliposomes. In the presence of the substrates (NaHCO₃ or Na₂CO₃) for the cotransporter, the inhibitory effect of the antibody on cotransporter activity was significantly blunted as compared with the inhibition observed in the absence of substrates. Western blot analysis of rabbit kidneys showed that the antibodies recognized strongly a 56 kDa protein band in microsomes of the inner stripe of outer medulla and inner medulla, but not in the outer stripe of outer medulla. A 56 kDa protein band was recognized in microsomes of the stomach, liver, esophagus, and small intestine but was not detected in red blood cell membranes. Localization of the Na⁺/HCO⁻ ₃ cotransporter protein by immunogold technique revealed specific labeling of the cotransporter on the basolateral membranes of the proximal tubules, but not in the brush border membranes. These results demonstrate that the polyclonal antibodies against the 56 kDa basolateral protein inhibit the activity of the Na⁺/HCO⁻ ₃ cotransporter suggesting that the 56 kDa protein represents the cotransporter or a component thereof. These antibodies interact at or near the substrate binding sites. The Na⁺/HCO cotransporter protein is expressed in different regions of the kidneys and in other tissues. Received: 27 January 1996/Revised: 23 July 1996     

A role for nucleoporin FG repeat domains in export of human immunodeficiency virus type 1 Rev protein and RNA from the nucleus.

The human immunodeficiency virus type 1 Rev protein contains a nuclear export signal (NES) that is required for Rev-mediated RNA export in mammals as well as in the yeast Saccharomyces cerevisiae. The Rev NES has been shown to specifically interact with a human (hRIP/RAB1) and a yeast (yRip1p) protein in the two-hybrid assay. Both of these interacting proteins are related to FG nucleoporins on the basis of the presence of typical repeat motifs. This paper shows that Rev is able to interact with multiple FG repeat-containing nucleoporins from both S. cerevisiae and mammals; moreover, the ability of Rev NES mutants to interact with these FG nucleoporins parallels the ability of the mutants to promote RNA export in yeast and mammalian cells. The data also show that, after Xenopus oocyte nuclear injection, several FG nucleoporin repeat domains inhibit the export of both Rev protein and U small nuclear RNAs, suggesting that these nucleoporins participate in Rev-mediated and cellular RNA export. Interestingly, not all FG nucleoporin repeat domains produced the same pattern of RNA export inhibition. The results suggest that Rev and cellular mediators of RNA export can interact with multiple components of the nuclear pore complex during transport, analogous to the proposed mode of action of the nuclear protein import receptor.