Changes in intracellular proteolysis in Escherichia coli during prolonged growth with a low concentration of dihydrostreptomycin

Abstract Escherichia coli was cultured with a low concentration of dihydrostreptomycin (2.5 μg/ml.). Growth was similar to untreated controls for 10 h after which a slow decline in growth rate occurred; growth ceased after 20 h. Intracellular catabolism of pulse-labelled protein synthesised at various points during the antibiotic treatment increased during the first 10 h, but during the second 10 h proteolysis progressively declined to almost control values. The production of an aberrant proteolytic system is one possible explanation.     

On the Stability of Messenger RNA and Ribosomal RNA in the Brains of Control Human Subjects and Patients with Alzheimer''s Disease

The levels of the mRNAs encoding the G protein subunits GS alpha, G beta 1, and G beta 2 were measured by northern blotting in the frontal cortex and hippocampus of control subjects and of patients with a clinical and histopathological diagnosis of dementia of the Alzheimer type (DAT). There was no significant difference, in either brain region, between the control and DAT groups for any of the G protein mRNAs measured. The degree of intersubject variability was very high, e.g., GS alpha mRNA in the frontal cortex (mean optical density +/- SD) was 405 +/- 342 in the control group versus 305 +/- 207 in the DAT group. The extent of generalised RNA degradation was assessed by detecting the breakdown products of 28S rRNA. RNA degradation was present in tissue samples from every human subject studied. The extent of 28S rRNA degradation in each subject was found to be related to the levels of G protein mRNA detected. The degree of RNA degradation in human subjects was found to be very variable and unaffected by the presence of DAT. RNA degradation correlated poorly with postmortem interval and this was confirmed by a controlled study of postmortem degradation in rat tissue. The possibility that the relative hypoxia and ischaemia in patients immediately before death could influence RNA degradation is discussed. The variable extent of RNA degradation means that great care must be taken to ensure the validity of RNA analyses undertaken in human postmortem brain, particularly when techniques are employed (such as in situ hybridisation) that themselves give no indication of RNA integrity.     

Comparison of properties of commercially available crystalline native and recombinant firefly luciferases

Commercially available crystalline native and recombinant firefly luciferases were compared. The two types of luciferase had indistinguishable responses to variation in ATP and luciferin concentrations and to omission of reaction components. The time courses of light production, the responses to nucleotide analogues, and the stability of the enzymes under several storage conditions were identical. The native enzyme had a slightly greater specific activity and was more sensitive to trypsin degradation. These differeces are probably attributable to differences in conformation.     

Kinetic studies of phenol degradation by Rhodococcus sp. P1 II. Continuous cultivation

The degradation of phenol by Rhodococcus sp. P1 was studied in continuous culture systems. The organism could be adapted by slowly increasing concentration, step by step, up to 30.0 g · 1^-1 phenol in the influent. The degradation rate reached values of about 0.3 g · g dry mass^-1 ·h^-1. Large step increases in phenol concentration and addition of further substrates (e.g., catechol) were tolerated up to a certain concentration. With increasing dilution rate and increasing inlet phenol concentration the stability of the system decreased.     

The relation between spruce monoculture and incipient podzolisation in ochreous brown earths of the Belgian Ardennes

Summary The effects of coniferous monoculture on the distribution of C, Fe and Al in the upper horizons of brown ochreous earths of the Belgian Ardennes were investigated by comparing 5 soil profiles developed under an 80 year old spruce stand with another 5 profiles developed under a climactic broad-leaved forest (beechwood).Organic carbon, Fe and Al were extracted with 0.1N NaOH/Na-tetraborate solution buffered at pH 9.7: recent studies have shown that this extraction is particularly appropriate for the detection of incipient podzolisation in brown earths-brown podzolic soils intergrades.Even if most of the classic podzolisation indexes fail to illustrate differences, nevertheless our results show that fulvic acids and organo-ferric complexes are present in significantly greater amounts in the upper part of the cambic (B)₁ horizon of the soils developed under conifers. Moreover, this podzolic tendancy is confirmed by the weathering patterns of the clay minerals in the A₁(B) horizons developed under spruces, i.e. a more pronouced weathering of chloritic layers than those observed in the beechwood soil, with a correlative genesis of more abundant smectite-like minerals. One may therefore conclude that the change in the humus type (moder to mor) after the planting of spruce trees, has been sufficient, within the local climatic and edaphic context, to promote incipient podzolisation.     

Studies on glucosinolate degradation in Lepidium sativum seed extracts

Analysis of Lepidium sativum seeds showed the presence of allyl, 2-phenethyl and benzyl glucosinolates, the first two being reported for the first time from this source. The effects of temperature, pH of the extraction medium and the length of time allowed for autolysis were assessed on the benzyl glucosinolate degradation products in seed extracts. In particulàr benzyl thiocyanate was not produced at higher temperatures but at ambient and lower temperatures it exceeded isothiocyanate. Nitrile was always the major product under the conditions studied, ever at pH levels as high as 7.4. Five new possible benzyl glucosinolate degradation products were detected and evidence is presented that benzaldehyde and benzyl alcohol could be secondary products formed thermally from isothocyanate and thiocyanate, respectively. Benzyl mercaptan and benzyl methyl sulphide also appear to be thermally produced.     

Synthesis and degradation of proteins during wheat endosperm development

Synthesis of proteins rich in lysine declines progressively with endosperm development and these proteins appear to be degraded preferentially at later stages. The proteolytic enzymes in extracts of endosperms at a late stage of development release considerably more lysine radioactivity from labelled endosperm proteins as compared with the enzymes in endosperms at an early stage.     

Binding of the erlin1/2 complex to the third intralumenal loop of IP3R1 triggers its ubiquitin-proteasomal degradation

Long-term activation of inositol 1,4,5-trisphosphate receptors (IP₃Rs) leads to their degradation by the ubiquitin–proteasome pathway. The first and rate-limiting step in this process is thought to be the association of conformationally active IP₃Rs with the erlin1/2 complex, an endoplasmic reticulum–located oligomer of erlin1 and erlin2 that recruits the E3 ubiquitin ligase RNF170, but the molecular determinants of this interaction remain unknown. Here, through mutation of IP₃R1, we show that the erlin1/2 complex interacts with the IP₃R1 intralumenal loop 3 (IL3), the loop between transmembrane (TM) helices 5 and 6, and in particular, with a region close to TM5, since mutation of amino acids D-2471 and R-2472 can specifically block erlin1/2 complex association. Surprisingly, we found that additional mutations in IL3 immediately adjacent to TM5 (e.g., D2465N) almost completely abolish IP₃R1 Ca²⁺ channel activity, indicating that the integrity of this region is critical to IP₃R1 function. Finally, we demonstrate that inhibition of the ubiquitin-activating enzyme UBE1 by the small-molecule inhibitor TAK-243 completely blocked IP₃R1 ubiquitination and degradation without altering erlin1/2 complex association, confirming that association of the erlin1/2 complex is the primary event that initiates IP₃R1 processing and that IP₃R1 ubiquitination mediates IP₃R1 degradation. Overall, these data localize the erlin1/2 complex–binding site on IP₃R1 to IL3 and show that the region immediately adjacent to TM5 is key to the events that facilitate channel opening.