A fast dynamic mode of the EF‐G‐bound ribosome

A key intermediate in translocation is an ‘unlocked state’ of the pre‐translocation ribosome in which the P‐site tRNA adopts the P/E hybrid state, the L1 stalk domain closes and ribosomal subunits adopt a ratcheted configuration. Here, through two‐ and three‐colour smFRET imaging from multiple structural perspectives, EF‐G is shown to accelerate structural and kinetic pathways in the ribosome, leading to this transition. The EF‐G‐bound ribosome remains highly dynamic in nature, wherein, the unlocked state is transiently and reversibly formed. The P/E hybrid state is energetically favoured, but exchange with the classical P/P configuration persists; the L1 stalk adopts a fast dynamic mode characterized by rapid cycles of closure and opening. These data support a model in which P/E hybrid state formation, L1 stalk closure and subunit ratcheting are loosely coupled, independent processes that must converge to achieve the unlocked state. The highly dynamic nature of these motions, and their sensitivity to conformational and compositional changes in the ribosome, suggests that regulating the formation of this intermediate may present an effective avenue for translational control.     

Studies on ulcerative dermal necrosis of salmonids

Fourteen fresh run salmon Satmo salar L. with early extant lesions of ulcerative dermal necrosis (UDN) were kept in separate tanks and treated with zinc free malachite green. Ten of the fish were held at 10° C and 4 at 2° C. The treatment precluded infection with Saprolegnia fungus and allowed natural resolution of the lesions. There was a marked difference in rate of healing between warm and cold water conditions.
Histological examination of healing lesions at different stages showed that there was a primary invasion of cuboidal epithelium over the collagen scar followed by a phase of disorganized proliferation which eventually organized itself into normal epithelium. Melanocytes were very obvious in the dermis of healing lesions.     

Translational control during the acute phase response. Ferritin synthesis in response to interleukin-1.

Interleukin-1 (IL-1 beta) increases the synthesis of both heavy and light (L)-ferritin subunits when added to human hepatoma cells (HepG2) grown in culture. RNase protection and Northern blot analysis with L-ferritin probes revealed that no changes in L-ferritin mRNA levels occur after cytokine stimulation. However, the induction coincides with an increased association of the L-subunit mRNA with polyribosomes. Since the recruitment of stored ferritin mRNA onto polyribosomes is seen when iron enters the cell, the effect of IL-1 beta on iron uptake was tested and was found to be unaffected by the lymphokine. Neither transferrin receptor mRNA levels nor the number of receptors displayed on the cell surface was affected by IL-1 beta. However, the action of the cytokine on ferritin translation is inhibited by the action of the intracellular iron chelator deferoxamine. These data indicate that IL-1 beta induces ferritin gene expression by translational control of its mRNA. The pathway of induction is different from iron-dependent ferritin gene expression whereas regulation requires the background presence of cellular iron.     

Identification of major common extracellular proteins secreted by Aeromonas salmonicida strains isolated from diseased fish.

Ten different strains of Aeromonas salmonicida that were isolated from diseased fish were grown under identical conditions (24 h at 25 degree C) in 3% (wt/vol) tryptone soya broth medium supplemented with vitamins and inorganic ions. In each case the extracellular proteins that were formed were compared by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and it was found that there were two significant common components, one with a molecular weight of 70,000 and the other with a weight of 56,000. Application of enzyme purification techniques to the supernatant fraction proteins of a culture of one of the strains resulted in the isolation of a 70-kilodalton (kDa) component, which was found to be a serine protease, and a 56-kDa component, which was hemolytic to trout erythrocytes. Rocket immunoelectrophoresis with rabbit antibodies to the isolated protease and hemolysin showed the same antigenic components in the supernatant fractions of all the cultures. These activities were assayed, and protease activity was found to vary by a factor of three, from 59 to 195 U/ml, while the range of hemolytic activity was over a narrow band, from 28 to 43 U/ml. There was an inconsistency between the immunoelectrophoretic and direct assay data in only one case. This indicated the presence of additional hemolytic activity, in addition to the 56-kDa component. The detection of large amounts of the same protease and hemolysin, two potent degradative activities, in a random series of strains of A. salmonicida suggests that they may be obligatory virulence factors in the development of furunculosis.     

The rabbit alpha-like globin gene cluster is polymorphic both in the sizes of BamHI fragments and in the numbers of duplicated sets of genes

The alpha-like globin gene cluster in rabbits contains embryonic zeta- globin genes, an adult alpha-globin gene, and theta-globin genes of undetermined function. The basic arrangement of genes, deduced from analysis of cloned DNA fragments, is 5'-zeta 0-zeta 1-alpha 1-theta 1- zeta 2-zeta 3-theta 2-3'. However, the pattern of restriction fragments containing zeta- and theta-globin genes varies among individual rabbits. Analysis of BamHI fragments of genomic DNA from 24 New Zealand white rabbits revealed eight different patterns of fragments containing zeta-globin genes. The large BamHI fragments containing genes zeta 0 and zeta 1 are polymorphic in length, whereas a 1.9-kb fragment containing the zeta 2 gene and the 3.5-kb fragment containing the zeta 3 gene do not vary in size. In contrast to this constancy in the size of the restriction fragments, the copy number of the zeta 2 and zeta 3 genes does vary among different rabbits. No length polymorphism was detected in the BamHI fragments containing the theta-globin genes, but again the copy number varies for restriction fragments containing the theta 2 gene. The alpha 1- and theta 1-globin genes are located in a nonpolymorphic 7.2-kb BamHI fragment. The combined data from hybridization with both zeta and theta probes shows that the BamHI cleavage pattern does not vary within the region 5'-alpha 1-theta 1- zeta 2-zeta 3-theta 2-3', but the pattern genomic blot-hybridization patterns for the progeny of parental rabbits with different zeta-globin gene patterns shows that the polymorphic patterns are inherited in a Mendelian fashion. Two different haplotypes have been mapped based on the genomic blot-hybridization data. The variation in the alpha-like globin gene cluster in the rabbit population results both from differences in the copy number of the duplication block containing the zeta-zeta-theta gene set and from the presence or absence of polymorphic BamHI sites.      

Effect of microorganisms on rate of liquid extraction of ethanol from fermentation broths

Liquid extraction is one means of removing metabolic products continuously during a fermentation and so reducing product inhibition. It is known that microbial organisms are attracted to liquid-liquid interfaces, and it is important for the design of extraction systems to establish if this has a detrimental effect on the rate of extraction. The extraction of ethanol from aqueous suspensions of yeast (Saccharomyces cerevisiae) using n- decanol is described in this paper. It was found that the presence of the yeast cells severely reduced the rate of ethanol extraction. The overall mass transfer coefficient was reduced from 5.0 x 10(-6) to 0.7 x 10(-6) m/s. This reduced overall mass transfer coefficient was unaffected by yeast concentration in the range 0.1-20 kg/m(3). The results are consistent with the yeast cells adsorbing to the interface in closely packed layers and preventing mass transfer by simply reducing the available interfacial area. Optical microscope observations confirmed that a yeast layer several cell diameters thick rapidly built up at the interface when a small decanol droplet was added to a yeast suspension.     

Trimethyloxonium modification of single batrachotoxin-activated sodium channels in planar bilayers. Changes in unit conductance and in block by saxitoxin and calcium

Single batrachotoxin-activated sodium channels from rat brain were modified by trimethyloxonium (TMO) after incorporation in planar lipid bilayers. TMO modification eliminated saxitoxin (STX) sensitivity, reduced the single channel conductance by 37%, and reduced calcium block of inward sodium currents. These effects always occurred concomitantly, in an all-or-none fashion. Calcium and STX protected sodium channels from TMO modification with potencies similar to their affinities for block. Calcium inhibited STX binding to rat brain membrane vesicles and relieved toxin block of channels in bilayers, apparently by competing with STX for the toxin binding site. These results suggest that toxins, permeant cations, and blocking cations can interact with a common site on the sodium channel near the extracellular surface. It is likely that permeant cations transiently bind to this superficial site, as the first of several steps in passing inward through the channel.