35S-β-glucuronidase gene blocks biological effects of cotransferred iaa genes

The iaaM and iaaH genes of Agrobacterium tumefaciens and Agrobacterium rhizogenes play an important role in crown gall and hairy root disease. The iaaM gene codes for tryptophan monooxygenase which converts tryptophan into indole-3-acetamide (IAM). IAM is converted into the auxin indole-3-acetic acid (IAA) by indoleacetamide hydrolase, encoded by the iaaH gene. In functional studies on the activity of the iaa genes of the TB region of the A. tumefaciens biotype III strain Tm4, the frequently used 35S--glucuronidase (35S-UidA or GUS) marker gene was found to inhibit IAA synthesis and root induction encoded by the TB iaa genes. To exert this inhibition, the 35S-UidA gene must be cotransferred with the iaaH gene. The 35S promoter alone is sufficient to cause the inhibitory effect.     

Frontiers in mycoplasma pathogenicity

Four different strains ofLactobacillus delbrueckii subsp.bulgaricus (Ss₁ and Yop₁₂) andStreptococcus salivarius subsp.thermophilus (Ss₂ and Yop₉) were isolated from two different yogurt sources in Argentina. In medium containing different carbon sources: lactose, fructose, sucrose or glucose plus fructose, the growth of a mixed culture (Yop₁₂+Ss₂) shows stimulation ofS. thermophilus and inhibition ofL. bulgaricus with respect to pure cultures. Both microorganisms in mixed culture grew less well on glucose plus galactose. However, in medium with glucose or galactose, both microorganisms were stimulated.     

Basic carboxyl groups of hemoglobin S: Influence of oxy-deoxy conformation on the chemical reactivity of Glu-43(β)

The -carboxyl groups of Glu-43() and Glu-22() of hemoglobin-S (HbS), two intermolecular contact residues of deoxy protein, are activated by carbodiimide atp H 6.0. The selectivity of the modification by the two nucleophiles, glycine ethyl ester (GEE) and glucosamine, is distinct. Influence ofN-hydroxysulfosuccinimide, a reagent that rescues carbodiimide-activated carboxyl (O-acyl isourea) as sulfo-NHS ester, on the overall selectivity and efficiency of the coupling of Glu-22() and Glu-43() with nucleophiles has been investigated. Sulfo-NHS increases the extent of coupling of nucleophiles to HbS. The rescuing efficiency of sulfo-NHS(increase in modification) with GEE and galactosamine as nucleophiles is 2.0 and 2.8, respectively. In the presence of sulfo-NHS, the extent of modification of a carboxyl group is a direct reflection of the extent to which it is activated (i.e., the protonation state of the carboxyl group). The modification reaction exhibits very high selectivity for Glu-43() with GEE and galactosamine (GA) in the presence of sulfo-NHS. From the studies of the kinetics of amidation of oxy-HbS at its Glu-43() (i.e., chemical reactivity) as a function of thepH in the region of 5.5–7.5, the apparentpKa of its -carboxyl group has been calculated to be 6.35. Deoxygenation of HbS, nearly doubles the chemical reactivity of Glu-43() of HbS atpH 7.0. It is suggested that the increased hydrophobicity of the microenvironment of Glu-43(), which occurs on deoxygenation of the protein, is reflected as the increased chemical reactivity of the -carboxyl group and could be one of the crucial preludes to the polymerization process.     

Dietary modulation of α-amylase activity in eight geographical strains of Sitophilus oryzae and Sitophilus zeamais

Rank transformation of specific activity values of -amylase across four strains of Sitophilus oryzae (L.) and four strains of S. zeamais Motschulsky indicates that levels of these predominant enzymes are highest in adults feeding on hulled barley or long-grain brown rice. Intermediate activity levels are found in weevils feeding on yellow corn (maize) and lowest levels are found in wheat-fed weevils. Although extracts prepared from barley contain inhibitory activity against two purified isoamylases from S. oryzae, levels of the naturally-occurring -amylase inhibitors against these two enzymes are about 2.2-fold and 6.1-fold, respectively, more concentrated in wheat. Ingestion of these amylase inhibitors and formation of an inactive enzyme:inhibitor complex with previously secreted amylase may account for the lower activity of amylase in weevils of both species feeding on wheat. Amylase levels across all strains feeding on a given diet are about 2-fold higher in S. oryzae than in S. zeamais. Significant differences in activity levels were also found between strains in both species. Since -amylase is a predominant digestive hydrolase in these species, the degree to which cereal diets affect amylase levels may indicate their suitability as potential hosts.

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Résumé La transformation de rang des valeur d'activité spécifique de l'-amylase de 4 souches de S. oryzae et de 4 souches de S. zeamais montre que les niveaux les plus élevés de ces enzymes prédominantes s'observent chez les adultes nourris d'orge mondé ou de riz brun á grains longs. Des niveaux intermédiaires d'activité ont été obtenus chez les insectes élevés sur maïs jaune, et les niveaux les plus faibles chez ceux élevés sur blé. Bien que les extraits préparés à partir d'orge présentent une activité inhibitrice de deux isoamylases purifiées de S. oryzae, les niveaux des inhibiteurs naturels -amylase de ces deux enzymes sont environ respectivement 2,2 et 6,1 fois plus concentrés dans le blé. L'ingestion de ces inhibiteurs d'amylase et la formation d'un complexe enzyme inactive/inhibiteur avec l'amylase secrétée antérieurement, peut rendre compte de la plus faible activité de l'amylase chez les charançons consommant du blé. Le niveau d'amylase de S. oryzae est 2 fois plus élevé que celui de S. zeamais pour toutes les souches élevées sur un régime donné. Des niveaux d'activité significativement différents ont été trouvés suivant les souches pour chacune des deux espèces. Puisque l'amylase est la principale hydrolase digestive de ces espèces, l'intensité de la modification des teneurs en amylase par la consommation de céréales peut indiquer leur adéquation comme hôtes potentiels.

     

A Nerve Growth Factor-Sensitive S6 Kinase in Cell-Free Extracts from PC 12 Cells

Soluble extracts from nerve growth factor (NGF)-stimulated PC12 cells prepared by alkaline lysis show a two- to 10-fold greater ability to phosphorylate the 40S ribosomal protein S6 than do extracts from control cells. The alkaline lysis method yields a preparation of much higher specific activity than does sonication. Half-maximal incorporation of 32P from [32P]ATP into S6 occurred after 4-7 min of NGF treatment. The partially purified NGF-sensitive S6 kinase has a molecular weight of 45,000. It is not inhibited by NaCl, chlorpromazine, or the specific inhibitor of cyclic AMP (cAMP)-dependent protein kinase, nor is it activated by addition of diolein plus phosphatidylserine. Trypsin treatment of either crude extracts or partially purified S6 kinase from control or NGF-treated cells was without effect. These data suggest that the S6 kinase stimulated by NGF is neither cAMP-dependent protein kinase or protein kinase C nor the result of tryptic activation of an inactive proenzyme. Treatment of intact cells with dibutyryl cAMP or 5'-N-ethylcarboxamideadenosine also increases the subsequent cell-free phosphorylation of S6. This observation suggests that cAMP-dependent protein kinase may be involved in the phosphorylation of S6 kinase.     

(6R)-Tetrahydrobiopterin Increases the Activity of Tryptophan Hydroxylase in Rat Raphe Slices

The effects of (6R)- and (6S)-tetrahydrobiopterin (BPH4), tetrahydroneopterin, and 6-methyltetrahydropterin on the activity of tryptophan hydroxylase were investigated in rat raphe slices. The activity of tryptophan hydroxylase was estimated by measurement of 5-hydroxytryptophan (5-HTP) formation under inhibition of aromatic L-amino acid decarboxylase with use of HPLC-fluorometric detection. (6R)-BPH4 (the naturally occurring form) at 42 microM, tetrahydroneopterin at 50 microM, and 6-methyltetrahydropterin at 100 microM increased tryptophan hydroxylase activity to 350, 145, and 146% of control values, respectively. (6S)-BPH4, however, had no significant effects on tryptophan hydroxylase activity. These results suggest that tryptophan hydroxylase is subsaturating in vivo for the naturally occurring cofactor, (6R)-BPH4, and that the concentration of (6R)-BPH4 may play an important role for the regulation of tryptophan hydroxylase activity in vivo.     

Critical nuclear DNA size and distribution associated with S phase initiation

Using HeLa S-3 cells synchronized by selective detachment, in this paper we report a parallel study of nuclear morphology and autoradiography grain patterns between middle G₁ and middle S phases: Our results show two distinct [³H]-thymidine labeling patterns. The first “peripheral” labeling pattern has a characteristic nuclear size distribution, in contrast to the heterogeneous and varying size distributions of Feulgen-stained nuclei, and apparently is characteristic of very early S phase. The sizes of the second labeling pattern—homogeneous or inhomogeneous grain distribution throughout the nucleus—are equal or larger than the first and vary with S phase progression. Together, the corresponding nuclear sizes of the labeled nuclei represent the larger extreme of nuclear areas, and the labeling index closely parallels the fraction of nuclei with areas larger than the minimum size of the labeled nuclei. These results suggest a characteristic nuclear size (reflecting unique intranuclear DNA distribution) as a necessary, if not sufficient, requirement for S phase initiation. Parallel experimentation with rat liver cells—synchronized in vivo by partial hepatectomy and analyzed by thin section autoradiography—confirms the existence of a peripheral labeling pattern in both the very early part and the very late part of S phase, which reconciles our data with previous results and points to the fact that both initiation and termination sites for DNA replication are near the nuclear periphery.     

The S-state dependence of Cl binding to plant Photosystem II

A combined single-turnover flash and ³⁵Cl NMR technique has been used to monitor S-state dependence of Cl⁻ binding to PS-II particles derived from mangrove (Avicennia marina). No detectable high-affinity binding was found to particles in the S₀ and S₁ states, but binding with an affinity comparable to that which activates O₂ evolution was found in the S₂ and S₃ states.     

An Escherichia coli mutant unable to support site-specific recombination of bacteriophage lambda

We report the isolation of mutations in, and the characterization of, an Escherichia coli gene, hip, that is required for site-specific recombination of phage lambda. hip mutants are recessive and are located near minute 20 on the linkage map. The gene product is not vital to bacterial growth, since deletion mutants are viable. The absence of hip product reduces lambda integration to barely detectable levels and also reduces prophage excision, but less drastically. Certain mutations in the lambda int gene partially restore integration and excision in hip- hosts. Homologous recombination promoted by recA does not require hip function. In addition to their defect in site-specific recombination, hip mutants are unable to support lytic growth of phage Mu or of certain lambda mutants. Their pleiotropic phenotype closely resembles that of himA mutants, but complementation, mapping and DNA sequencing show that hip and himA are different genes.     

What are mycoplasmas: The relationship of tempo and mode in bacterial evolution

Summary In phenotype the mycoplasmas are very different from ordinary bacteria. However, genotypically (i.e., phylogenetically) they are not. On the basis of ribosomal RNA homologies the mycoplasmas belong with the clostridia, and indeed havespecific clostridial relatives. Mycoplasmas are, however, unlike almost all other bacteria in the evolutionary characteristics of their ribosomal RNAs. These RNAs contain relatively few of the highly conserved oligonucleotide sequences characteristic of normal eubacterial ribosomal RNAs. This is interpreted to be a reflection of an elevated mutation rate in mycoplasma lines of descent. A general consequence of this would be that the variation associated with a mycoplasma population is augmented both in number and kind, which in turn would lead to an unusual evolutionary course, one unique in all respects. Mycoplasmas, then, are actually tachytelic bacteria. The unusual evolutionary characteristics of their ribosomal RNAs are the imprints of their rapid evolution.