The synthesis of β-galactosidase inEscherichia coli in the presence of 7-azatryptophan

The presence of 7-azatryptophan an analogue of tryptophan in the growth medium ofEscherichia coli resulted in a considerable inhibition of the synthesis of active β-galactosidase. No synthesis of an immunologically cross-reacting protein was detected. In addition, the replacement of tryptophan by the analogue rendered the enzyme more susceptible to heat, urea and trypsin as compared with the normal enzyme. The inhibition of growth and enzyme synthesis by 7-azatryptophan was reversed by tryptophan. The analogue did not exhibit any effect on the synthesis and activity of β-galactoside permease.     

Possible Effect of Activation of α7-Nicotinic Acetylcholine Receptors in the Mitochondrial Membrane on the Development of Apoptosis

Using flow cytometry and sandwich-immunoenzyme assay, we showed that nicotinic acetylcholine receptors with a subunit α7 (nAChRs α7) expressed in the outer mitochondrial membrane are involved in the control of mitochondria-dependent apoptosis. Pre-incubation of the mitochondria with an nAChRs α7 agonist, choline, decreased dissipation of the membrane potential of these organelles induced by the action of 0.5 mM hydrogen peroxide (H₂O₂) but did not influence the analogous effect of a high Ca²⁺ concentration (90 μM). Agonists of nAChRs α7 (choline, acetylcholine, and PNU 282987), or an inhibitor of voltage-dependent anion channels, DIDS, prevented the release of cytochrome c from the intermembrane mitochondrial space under the action of H₂O₂. In contrast, an antagonist of nAChRs α7, methyllycaconitine, promoted the release of cytochrome c and prevented the effects of agonists. The obtained data confirm the active involvement of nAChRs α7 and voltage-dependent anion channels in the process of formation of mitochondrial pores. In this case, agonists of mitochondrial nAChRs α7 subunits exert an antiapoptotic effect, while antagonists of mitochondrial nAChRs α7 subunits manifest a proapoptotic action.     

Immunocytochemical localization and concentrations of the α and γ subunits of 7S-nerve growth factor in the submandibular gland of the mouse

Summary For unexplained reasons, nerve growth factor (NGF) exists in very high concentrations in the submandibular gland of the mouse. The NGF in the gland, called 7S-NGF, is a non-covalent complex of three protein subunits, named -, - and -NGF. All the known biological activity resides in the -NGF subunit, and previous studies have shown that -NGF is present in much greater concentrations in the male submandibular gland than in the female gland. The higher concentration in the male is due to the fact that -NGF is synthesized in the granular tubule cells of the submandibular gland. These cells are much more numerous in the male gland.In contrast to -NGF, neither the concentrations of and subunits nor their cellular localization in the mouse submandibular gland have been established. In this study, radioimmunoassays specific for . and subunits determined that both are present in much higher concentrations in the male gland. Immunocytochemical work localized both subunits in the granular tubule cell in the male and female submandibular gland. This indicates that all the components of 7S-NGF exist in a single cell type in the gland and suggests that 7S-NGF can be formed within this cell and secreted as a complex into the saliva.     

Induction of antibodies to particular sites of the α7 subunit of the nicotinic acetylcholine receptor in mice of different lines

Five synthetic fragments of the N-terminal domain of the α7 subunit of the human nicotinic acetylcholine receptor (α7 nAChR) that correspond to theoretically calculated B epitopes and T helper epitopes of the protein and contain from 16 to 29 amino acid residues were tested for the ability to stimulate the formation of antibodies in mice of three lines having H-2^d, H-2^b, and H-2^k haplotypes of the major histocompatibility complex. It was shown that, in the free (unconjugated) form, all the peptides stimulate the formation of antibodies at least in one mouse line. Most of the peptides induced the formation of antibodies in BALB/c mice (haplotype H-2^d); therefore, more detailed studies were carried out on these animals. The free peptides and/or their conjugates with keyhole limpet hemocyanin were demonstrated to be capable of stimulating the formation in BALB/c mice of antibodies that bind to the recombinant extracellular N-terminal domain of (α7 nAChRα. The epitope mapping of antipeptide antibodies carried out using truncated fragments helped reveal antipeptide antibodies to four regions of the α7 subunit: 1–23, 98–106, 159–168, and 173–188 (or 179–188).     

Alterations in the expression of the Atp7a gene in the early postnatal development of the mosaic mutant mice (Atp7amo-ms) – An animal model for Menkes disease

Copper is a trace element that is essential for the normal growth and development of all living organisms. In mammals, the ATP7A Cu-transporting ATPase is a key protein that is required for the maintenance of copper homeostasis. In both humans and mice, the ATP7A protein is coded by the X-linked ATP7A/Atp7a gene. Disturbances in copper metabolism caused by mutations in the ATP7A/Atp7a gene lead to severe metabolic syndromes Menkes disease in humans and the lethal mottled phenotype in mice. Mosaic is one of numerous mottled mutations and may serve as a model for a severe Menkes disease variant. In Menkes patients, mutations in the ATP7A gene often result in a decreased level of the normal ATP7A protein. The aim of this study was to analyse the expression of the Atp7a gene in mosaic mutants in early postnatal development, a critical period for starting copper supplementation therapy in both Menkes patients and mutant mice. Using real-time quantitative RT-PCR, we analysed the expression of the Atp7a gene in the brain, kidney and liver of newborn (P0.5) and suckling (P14) mice. Our results indicate that in mosaic P0.5 mutants, the Atp7a mRNA level is decreased in all analysed organs in comparison with wild-type animals. In two week-old mutants, a significant decrease was observed only in the kidney. In contrast, their hepatic level of Atp7a tended to be higher than in wild-type mice. We speculate that disturbance in the expression of the Atp7a gene and, consequently, change in the copper concentration of the organs, may contribute to the early fatal outcome of mosaic males.     

Identification of the 7S globulin with β-conglycinin in soybean seeds

The 7S globulin, a major ultracentrifugal component with the 11S globulin, was identical with β-conglycinin one of four antigenic components in the reserve proteins of soybean seeds (Glycine max). Double gel immunodiffusion and immunoelectrophoresis in agar gel were used for their identification. In addition, some characteristic properties on ultracentrifugation and in carbohydrate content agreed well between the proteins. Their MWs were ca 180000.     

Basic Characteristics of Maturity of the Brain Bioelectric Activity in Children 1–7 Years of Age

Electroencephalographic (EEG) examination of healthy children within the ages of one to seven years was conducted to determine the basic EEG characteristics of maturation of the brain bioelectric activity. It was found that the index and frequency of the activity, the average range of the rhythm assimilation reaction, and the expression of area-specific differences in the -rhythm and activation response are characterized by an increasing pattern of age-related changes in healthy children 1–7 years old. On the other hand, the index of the -rhythm showed a decreasing pattern of age-related changes as did the maximum (5–8) number of correlation relationships between these EEG measures.     

The pivotal role of the β7 strand in the intersubunit contacts of different human small heat shock proteins

Human αB-crystallin and small heat shock proteins HspB6 and HspB8 were mutated so that all endogenous Cys residues were replaced by Ser and the single Cys residue was inserted in a position homologous to that of Cys137 of human HspB1, i.e. in a position presumably located in the central part of β7 strand of the α-crystallin domain. The secondary, tertiary, and quaternary structures of thus obtained Cys-mutants as well as their chaperone-like activity were similar to those of their wild-type counterparts. Mild oxidation of Cys-mutants leads to formation of disulfide bond crosslinking neighboring monomers thus indicating participation of the β7 strand in intersubunit interaction. Oxidation weakly affects the secondary and tertiary structure, does not affect the quaternary structure of αB-crystallin and HspB6, and shifts equilibrium between monomer and dimer of HspB8 towards dimer formation. It is concluded that the β7 strand participates in the intersubunit interaction of four human small heat shock proteins (αB-crystallin, HspB1, HspB6, HspB8) having different structure of β2 strand of α-crystallin domain and different length and composition of variable N- and C-terminal tails.     

Influence of ampicillin,clindamycin or metronidazole upon the 7α-dehydroxylation of bile acids in the human intestine

The effect of ampicillin, clindamycin and metronidazole upon the 7-dehydroxylation of cholic acid by the microflora in the intestine was tested in healthy subjects. Clindamycin gave a significant decrease in the 7-dehydroxylation. With the other two antibiotics, no significant changes were seen.     

Immunocytochemical localization of a novel pituitary polypeptide “7B2” in the gastro-intestinal tract of the rat

Summary Immunoreactivity to the polypeptide designated 7B2 recently isolated from human and porcine pituitary glands, appears to be consistently confined to neuroendocrine and endocrine cells in various tissues. In rat gut, immunoreactive 7B2 was found in endocrine-paracrine cells. Highly labeled cells were found in the antrum of the stomach and, cells with lower concentrations, in the fundus, duodenum, jejunum and ileum. Except for a few cells which were simultaneously positive for 5-hydroxytryptamine, and a few others showing Grimelius's reaction, 7B2 cells do not exhibit argentaffin and/or argyrophil character. The 7B2 polypeptide seems to be distributed amongst several different types of endocrine cells in the gut.     

The stereochemistry of the hydrogen elimination in the biological conversion of cholest-7-en-3β-ol into cholesterol

1. The syntheses of Δ⁷-[4-¹⁴C]cholestenol (XVI, Scheme 3) and Δ⁷-[6α-³H]-cholestenol (XII, Scheme 2) are described. 2. The metabolism of doubly labelled Δ⁷-cholestenol (II, Scheme 1) by rat-liver homogenates was studied. 3. During the enzymic conversion of Δ⁷-cholestenol into cholesterol (IV, Scheme 1) the 6α-hydrogen atom of the former is lost and the overall reaction corresponds to a cis-elimination. 4. In the light of these results various mechanisms for the conversion of Δ⁷-cholestenol into cholesterol are discussed.     

Effect of 7-methylguanosine-5′-phosphate on the translation of rat milk-protein mRNAs in the wheat-germ cell-free system

The translation of polyadenylated and of non-poly-adenylated RNA obtained from lactating rat mammary gland was almost totally inhibited by 0,5 mM 7-methylguanosine-5-phosphate in the wheat-germ cell-free system, This inhibition was maintained during the preparation of the 9S whey-protein mRNA and of the 12S and ISS casein mRNAs, Chemical decapping of these mRNAs caused a similar reduction of their activity . Although a large fraction of milk-protein mRNAs have been reported to lack 3-polyadenylation, these results show that the mRNAs in the mammary gland do contain a 5-terminal 7-methylguanosine cap.     

Role of Protein Tyrosine Phosphatase Non-Receptor Type 7 in the Regulation of TNF-α Production in RAW 264.7 Macrophages

Protein tyrosine phosphatases play key roles in a diverse range of cellular processes such as differentiation, cell proliferation, apoptosis, immunological signaling, and cytoskeletal function. Protein tyrosine phosphatase non-receptor type 7 (PTPN7), a member of the phosphatase family, specifically inactivates mitogen-activated protein kinases (MAPKs). Here, we report that PTPN7 acts as a regulator of pro-inflammatory TNF-α production in RAW 264.7 cells that are stimulated with lipopolysaccharide (LPS) that acts as an endotoxin and elicits strong immune responses in animals. Stimulation of RAW 264.7 cells with LPS leads to a transient decrease in the levels of PTPN7 mRNA and protein. The overexpression of PTPN7 inhibits LPS-stimulated production of TNF-α. In addition, small interfering RNA (siRNA) analysis showed that knock-down of PTPN7 in RAW 264.7 cells increased TNF-α production. PTPN7 has a negative regulatory function to extracellular signal regulated kinase 1/2 (ERK1/2) and p38 that increase LPS-induced TNF-α production in macrophages. Thus, our data presents PTPN7 as a negative regulator of TNF-α expression and the inflammatory response in macrophages.     

Variation in the cell cycle time in the crypts of lieberkühn of the mouse

The durations of the phases of the cell cycle were measured at different levels in the jejunal crypts of male Balb/c mice. A mean cell cycle time of 12.3 h was found for the whole crypt. In cell positions 1 and 2, the cell cycle time was 16.7 h, and this time steadily decreased to a value of between 10 and 11 h for cell positions above 11. It is concluded that basally situated crypt cells in the mouse are cycling relatively slowly, and that they form the functional stem cell pool for the crypt. These cells may also compose the potential stem cell pool which repopulates the crypt after death of proliferative cells.     

Immunohistochemical localization of the angiotensin-(1–7) receptor Mas in the murine forebrain

Apart from the well-known biologically active angiotensin II, other biologically active angiotensins have been discovered, including angiotensin IV and angiotensin-(1–7). Some years ago, we and others discovered that the Mas proto-oncogene encodes a receptor that is essential for angiotensin-(1–7) signaling. Angiotensin-(1–7) is not only expressed in the periphery but also within the brain. Based on that, we examined the distribution of Mas within the murine brain, using an antibody directed against the 3^rd cytoplasmic loop of the receptor protein. Strongest Mas protein expression was detected in the dentate gyrus of the hippocampus and within the piriform cortex. However, Mas protein expression is not restricted to these areas, since Mas immunopositive neurons were also seen in different parts of the cortex, hippocampus, amygdala, basal ganglia, thalamus and hypothalamus. Based on the expression of Mas protein in the cortex and the limbic system, angiotensin-(1–7) signaling may play a role in synaptic plasticity, learning, memory and emotion, as has been described for angiotensin II and IV.