2 A prebiotic RNA apparatus functions within the contemporary ribosome

Ribosomes, the universal cellular machines, possess spectacular architecture accompanied by inherent mobility, allowing for their smooth performance as polymerases that translate the genetic code into proteins. The site for peptide bond formation is located within a universal internal semi-symmetrical region, which was identified within all contemporary ribosomes. The high conservation of this region implies its existence irrespective of environmental conditions and indicates that it may represent an ancient RNA molecular apparatus. Hence, we named it the “proto-ribosome”. This prebiotic pocket-like RNA entity is suggested to be capable to accommodate substrates whose stereochemistry enables the creation of chemical bonds. It could have evolved from an earlier catalytic RNA entity that we named the “pre-proto-ribosome”, presumed to be a molecular machine capable of performing various essential tasks in the RNA world, which was snatched by the amino acid invaders for producing proteins.     

Autoradiographic Analysis of [3H]Imipramine Binding in the Human Brain Postmortem: Effects of Age and Alcohol

In vitro quantitative autoradiography of high-affinity [3H]imipramine binding sites was performed on 16 human brains postmortem. The densities of binding sites were highest in the hypothalamus. Next, in descending order, were the basal and lateral nuclei of the amygdala; substantia innominata; insular cortex; the central nucleus of the amygdala; the anterior nucleus of the thalamus; the head of the caudate nucleus; portions of the frontal, parietal, and temporal cortex; claustrum; the granular layer of the dentate gyrus; substantia nigra; the pyramidal layer of CA fields; globus pallidus; red nucleus; and white matter. Imipramine binding was found to increase with age in a region-specific manner. The presence of alcohol had a similar effect, which was most pronounced in the hippocampus. Sex and time from death to autopsy did not affect imipramine binding, in our sample.     

Effect of Chronic Desipramine Treatment on Dihydroalprenolol, Imipramine, and Desipramine Binding Sites: A Quantitative Autoradiographic Study in the Rat Brain

Desmethylimipramine (DMI) administered once daily for 10 days caused a significant decrease in beta-adrenergic receptor binding, as measured by quantitative autoradiography in discrete brain regions. The decrease was observed 72 h after the last injection throughout the cortex and in hippocampus but not in other regions, much richer in beta-receptors, such as the caudate, olfactory tubercle, superior colliculus, dorsomedial thalamus, substantia nigra, or pineal. The same paradigm did not affect imipramine (IMI) binding in the cortex or in regions with high concentrations of IMI binding sites. DMI binding was not decreased, either. Significant increases in DMI binding were observed in frontal cortex and in the ventral aspect of the bed nucleus of the stria terminalis. We conclude that a reduction in tricyclic binding is not a general phenomenon following chronic treatment with tricyclic antidepressants, and changes in binding, when they do occur, are not correlated with areas of high binding site density.     

On peptide bond formation, translocation, nascent protein progression and the regulatory properties of ribosomes. Derived on 20 October 2002 at the 28th FEBS Meeting in Istanbul.

High-resolution crystal structures of large ribosomal subunits from Deinococcus radiodurans complexed with tRNA-mimics indicate that precise substrate positioning, mandatory for efficient protein biosynthesis with no further conformational rearrangements, is governed by remote interactions of the tRNA helical features. Based on the peptidyl transferase center (PTC) architecture, on the placement of tRNA mimics, and on the existence of a two-fold related region consisting of about 180 nucleotides of the 23S RNA, we proposed a unified mechanism integrating peptide bond formation, A-to-P site translocation, and the entrance of the nascent protein into its exit tunnel. This mechanism implies sovereign, albeit correlated, motions of the tRNA termini and includes a spiral rotation of the A-site tRNA-3' end around a local two-fold rotation axis, identified within the PTC. PTC features, ensuring the precise orientation required for the A-site nucleophilic attack on the P-site carbonyl-carbon, guide these motions. Solvent mediated hydrogen transfer appears to facilitate peptide bond formation in conjunction with the spiral rotation. The detection of similar two-fold symmetry-related regions in all known structures of the large ribosomal subunit, indicate the universality of this mechanism, and emphasizes the significance of the ribosomal template for the precise alignment of the substrates as well as for accurate and efficient translocation. The symmetry-related region may also be involved in regulatory tasks, such as signal transmission between the ribosomal features facilitating the entrance and the release of the tRNA molecules. The protein exit tunnel is an additional feature that has a role in cellular regulation. We showed by crystallographic methods that this tunnel is capable of undergoing conformational oscillations and correlated the tunnel mobility with sequence discrimination, gating and intracellular regulation.     

The maintenance ofBdellovibrio at low prey density

A mathematical model for the interaction ofBdellovibrio and its prey predicted that a relatively high prey density (7×10⁵ cells ml^–1) would be required for the establishment of an equilibrium in a mixed population [8]. The present report shows thatBdellovibrio can be maintained in a continuous culture when the prey cell density is much lower (2–5×10⁴ cells ml^–1), and closer to that of naturally occurring bacterial populations in sea waters.     

A Unique Ascorbate Peroxidase Active Component in the Cyanobacterium Synechococcus PCC 7942 (R2)

Ascorbate peroxidase active component (APAC) was purified and characterized in Synechococcus PCC 9742 (R2) cells. APAC was isolated from freshly harvested cells, by ion exchange chromatography on DEAE cellulose, ultrafiltration through a 3000 dalton cut off filter and high pressure liquid chromatography through a reversed phase C-18 column. APAC was found to be extremely stable to harsh treatments of boiling water for 30 min, acidification to pH 2.0 and proteolytic digestion. A close correlation between activity and iron content of APAC was observed throughout the purification steps. E.S.R. spectrum of APAC showed a resonance line at g = 4.3 in the oxidized from. Peroxide reduction by ascorbate decreased the E.S.R. signal, which reappeared upon reoxidation by H₂O₂. The affinities of APAC to H₂O₂ and ascorbate were high (0.38 mM and 0.2 mM, respectively). Amino acid composition analysis of APAC revealed the presence of glutamic acid: glycine: cysteine residues at 2: 1: 1 ratio.     

Unique properties of the camel erythrocyte membrane: II. Organization of membrane proteins

Camel erythrocyte membranes are distinguished by some unique properties of stability and composition. Notable is their abundance in proteins (protein: lipid ratio of 3 : 1). Membrane proteins of camel erythrocytes were compared with those of human erythrocytes, which have been intensively investigated. Proteins were extracted with various aqueous media (EDTA, alkaline or high ionic strength) and with ionic and non-ionic detergents and were analyzed by gel electrophoresis. In membranes of camel erythrocytes, the peripheral proteins constitute, proportionally, a much smaller fraction of total proteins than in the human erythrocyte, while their distribution is identical per unit of surface area. The camel erythrocyte membrane is particularly rich in integral proteins and in intramembranous particles. The proteins in this membrane are more closely organized than in the human system, as revealed by crosslinking and freeze-etching studies. It is proposed that protein-protein interaction of integral proteins, presumably constituting an “integral skeleton”, is a dominant structural feature stabilizing the camel erythrocyte membrane.     

Ribonuclease S-peptide. A model for molecular recognition.

The relationship of structure to function in the recognition of ribonuclease S-peptide by S-protein was studied by several methods. Liquid phase peptide synthesis was employed to generate analogs of S-peptide in which from 1 to 8 residues were deleted from the NH2-terminal end of the S-peptide. Additional derivatives were made by substitutions in the NH2-terminal three amino acids or by modifying the S-peptide analogs by trifluoroacetylation. The analogs were generated in the following way. S-Peptide was cleaved with chymotrypsin. The fragment obtained, RNase(9-20), was purified and lengthened step by step using liquid phase peptide synthesis. A second set of analogs were prepared by cleavage of CF3CO-S-peptide with elastase and the resulting CF3CO-RNase(7-20), similarly lengthened. The various analogs of S-peptide were tested in their capacity to combine with S-protein and regenerate biological activity as measured by Vmax and Kb. This work shows a positive contribution of every one of the first 8 NH2-terminal residues of S-peptide to the molecular recognition of S-protein in the presence of RNA substrate. Substitution of the first 3 residues by alanine or blocking of the free amino groups decreases recognition, indicating that the original primary structure is the most favorable one.     

Recent update on lactic acid bacteria producing riboflavin and folates: application for food fortification and treatment of intestinal inflammation

Lactic acid bacteria (LAB), widely used as starter cultures for the fermentation of a large variety of food, can improve the safety, shelf life, nutritional value and overall quality of the fermented products. In this regard, the selection of strains delivering health-promoting compounds is now the main objective of many researchers. Although most LAB are auxotrophic for several vitamins, it is known that certain strains have the capability to synthesize B-group vitamins. This is an important property since humans cannot synthesize most vitamins, and these could be obtained by consuming LAB fermented foods. This review discusses the use of LAB as an alternative to fortification by the chemical synthesis to increase riboflavin and folate concentrations in food. Moreover, it provides an overview of the recent applications of vitamin-producing LAB with anti-inflammatory/antioxidant activities against gastrointestinal tract inflammation. This review shows the potential uses of riboflavin and folates producing LAB for the biofortification of food, as therapeutics against intestinal pathologies and to complement anti-inflammatory/anti-neoplastic treatments.