Highly conserved 5S ribosomal RNA sequences in four rust fungi and atypical 5S rRNA secondary structure in Microstroma juglandis.

The 5S ribosomal RNA nucleotide sequences of five basidiomycetous fungi, Coleosporium tussilaginis , Gymnosporangium clavariaeforme , Puccinia poarum , Endophyllum sempervivi and Microstroma juglandis were determined. Despite high differentiation in their host spectra the four rust species are highly conserved with respect to their 5S rRna sequences, which fit with the basidiomycete cluster 5 described by Walker and Doolittle (1). The sequences obtained from the first three rust fungi were proven to be identical while the sequence from Endophyllum sempervivi showed two base substitutions compared with the other rust fungi. The Microstroma juglandis 5S rRNA sequence differs from all other basidiomycete 5S rRNA sequences published so far in respect to its secondary structure which shows an atypical 'CCA' loop in helix D, but it reveals typical basidiomycetous signature nucleotides. Therefore Microstroma juglandis represents a cluster of its own within the Basidiomycetes. A dendrogram was constructed based on Kimura's "Neutral Theory of Molecular Evolution".     

Energy-dependent inactivation of citrate lyase in Enterobacter aerogenes.

Enterobacter aerogenes was grown in continous culture with ammonia as the growth-limiting substrate, and changes in citrate lyase and citrate synthase activities were monitored after growth shifts from anaerobic growth on citrate to aerobic growth on citrate, aerobic growth on glucose, anaerobic growth on glucose, and anaerobic growth on glucose plus nitrate. Citrate lyase was inactivated during aerobic growth on glucose and during anaerobic growth with glucose plus nitrate. Inactivation did not occur during anaerobic growth on glucose, and as a result of the simultaneous presence of citrate lyase and citrate synthase, growth difficulties were observed. Citrate lyase inactivation consisted of deacetylation of the enzyme. The corresponding deacetylase could not be demonstrated in cell extracts, and it is concluded that, as in a number of other inactivations, electron transport to oxygen or nitrate was required for inactivation.     

Complexity of RNA in Eggs of DROSOPHILA MELANOGASTER and MUSCA DOMESTICA

Comparative measurements are presented of the sequence complexity of the RNA stored in the eggs of two dipteran flies, Musca domestica and Drosophila melanogaster. The genome of Musca is about five times the size of the Drosophila genome and contains about 3.6 times as much single-copy sequence. As shown earlier, the interspersion of repetitive and single-copy sequence is of the short-period form in Musca, and is of the long-period form in Drosophila. The egg RNA complexities were determined by hybridization of excess RNA with radioactively labeled single-copy DNA. Complexity is expressed as the length (in nucleotides) of diverse single-copy sequence represented in the RNA. The complexity of the RNA of the Musca egg is about 2.4 x 10⁷ nucleotides, and that of the Drosophila egg is about 1.2 x 10⁷ nucleotides. The RNA of the Musca egg is similar to or very slightly lower in complexity than that of other egg RNAs, e.g., those of Xenopus and sea urchin. Compared to all previously measured egg RNAs, Drosophila egg RNA is low in sequence complexity.     

Lectin-mediated sperm agglutination of bufo arenarum and leptodactylus chaquensis spermatozoa

Various plant lecins were employed in cell agglutination experiments to ascertain the presence of specific saccharides in the surface of B arenarum and L chaquensis spermatozoa. B arenarum spermatozoa were specifically agglutinated with Concanavalin A (Con A), phytohemagglutinin P (PHA-P), and wheat germ agglutinin (WGA), but not with soybean agglutinin (SBA). In contrast, L chaquensis spermatozoa were strongly agglutinated by SBA, WGA, and PHA-P. L chaquensis spermatozoa did not agglutinate with Con A even at high concentrations. Lectinmediated sperm agglutination was inhibited in the presence of specific lectinbinding sugars. Spermatozoa from both species were agglutinated randomly with all lectins suggesting a uniform distribution in the sperm surface of the lectinbinding saccharide ligands. B arenarum sperm agglutination induced by Con A is sensitive to temperature. B arenarum spermatozoa are more agglutinable at 24°C than at 4°C. These results suggest that lectin-binding site mobility is necessary for sperm agglutination.     

A sodium-stimulated ATP synthase in the acetogenic bacterium Acetobacterium woodii.

Experiments with resting cells of Acetobacterium woodii were performed to elucidate the coupling ion used by the ATP synthase. A. woodii synthesized ATP in response to an artificial delta pH, indicating the presence of a proton-translocating ATPase. On the other hand, a delta pNa, as well as a proton diffusion potential, could serve as a driving force for ATP synthesis with the latter strictly dependent on Na+. These results are indicative for the presence of a Na(+)-translocating ATP synthase in A. woodii.     

N5-methyl-tetrahydromethanopterin:coenzyme M methyltransferase of Methanosarcina strain Gö1 is an Na(+)-translocating membrane protein.

To determine the cellular localization of components of the methyltransferase system, we separated cell extracts of Methanosarcina strain G?1 into cytoplasmic and inverted-vesicle fractions. Measurements demonstrated that 83% of the methylene-tetrahydromethanopterin reductase activity resided in the cytoplasm whereas 88% of the methyl-tetrahydromethanopterin:coenzyme M methyltransferase (methyltransferase) was associated with the vesicles. The activity of the methyltransferase was stimulated 4.6-fold by ATP and 10-fold by ATP plus a reducing agent [e.g., Ti(III)]. In addition, methyltransferase activity depended on the presence of Na+ (apparent Km = 0.7 mM) and Na+ was pumped into the lumen of the vesicles in the course of methyl transfer from methyl-tetrahydromethanopterin not only to coenzyme M but also to hydroxycobalamin. Both methyl transfer reactions were inhibited by 1-iodopropane and reconstituted by illumination. A model for the methyl transfer reactions is presented.     

Effects of promoters on the enhancement of pectin methyl esterase expression inAspergillus niger

Summary A pectin methylesterase-encoding gene (pmeA)_has been cloned and transformed intoA. niger wild-type NRRL3. Transformants produced 20-fold more PME than the host strain. For studying the effects of different promoters on thepmeA expression two novel plasmids were constructed, in which thepmeA promoter was replaced by efficient promoters such as theA. nidulans glyceraldehyde-3-phosphate dehydrogenase (pK45) or theA. oryzae -amylase (pK61) promoter. The highest level of PME expression was achieved with theA. oryzae -amylase promoter, reaching a 200-fold increase compared to the production by the host strain.     

Authenticity and reconstitution of immobilized enzymes: characterization and denaturation/renaturation of glucoamylase II.

Glucoamylase II (GA II) immobilized to Eupergit C and CIZ as a porous and nonporous matrix shows enzymatic characteristics indistinguishable from those of the free enzyme, except for reduced specific activity. Since this decrease is equally observed for both matrices, it has to be ascribed to nonproductive fixation of the enzyme or steric hindrance rather that perturbations caused by "inner diffusion" effects. Authenticity refers to the optimum pH for catalytic activity, Michaelis constants for starch and maltoheptaose, as well as identical stability toward temperature, pH, and guanidinium chloride (GdmCl). On the basis of these data, the two-state mechanism observed for the equilibrium transitions of the free enzyme may be assumed to hold also for the immobilized enzyme. Renaturation after preceding denaturation in 6.4 and 7 M GdmCl leads to widely differing yields depending on the conditions. Shifting the denaturant concentration stepwise back to nondenaturing GdmCl concentrations leads to a broad range of "hysteresis" accompanied by aggregation. Rapid dilution of the free and immobilized enzymes at pH greater than 6 and sufficiently low protein concentration leads to reactivation yields of 80 and 45%, respectively. For the free enzyme, reconstitution at lower pH is determined by the kinetic competition of folding and aggregation. In the case of the immobilized enzyme, "entangling" of the matrix with the unfolded polypeptide chain competes with renaturation.     

AMONG- AND WITHIN-FLOWER COMPARISONS OF POLLEN TUBE GROWTH FOLLOWING SELF- AND CROSS-POLLINATIONS IN DIANTHUS CHINENSIS (CARYOPHYLLACEAE)

We compared the rate of pollen tube growth following self- and cross-pollinations both among and within flowers of two clones of Dianthus chinensis L. For among-flower comparisons, both styles of a flower were pollinated with either self- or cross-pollen. Within-flower comparisons were made between the two styles of the same flower, one of which was self-pollinated and the other cross-pollinated. Comparisons between flowers indicated that self-pollen grew slower than cross-pollen in both clones. However, differences in the growth rate of pollen tubes from self- and cross-pollinations were greater when comparisons were made between the two styles of the same flower than when pollinations were made in different flowers. These results suggest the existence of interstyle interactions in pollen tube growth.