The Endoplasmic Reticulum of Mung Bean Cotyledons: ROLE IN THE ACCUMULATION OF HYDROLASES IN PROTEIN BODIES DURING SEEDLING GROWTH

The subcellular localization of two hydrolases (ribonuclease and vicilin peptidohydrolase) which are synthesized de novo in the cotyledons of mung bean seedlings was studied. Earlier experiments had shown that both enzymes accumulate in the protein bodies in the course of seedling growth. Two methods to fractionate subcellular organelles were used to demonstrate that a significant proportion of the enzymes is organelle-associated. This proportion is highest (up to 50% for vicilin peptidohydrolase and 15% for ribonuclease) when synthesis of the enzymes has just started. Evidence obtained with isopycnic sucrose gradients indicates that both hydrolases are associated with membranes rich in NADH-cytochrome c reductase, a marker enzyme for the endoplasmic reticulum (ER). The hydrolases band with the NADH-cytochrome c reductase under conditions where the ribosomes remain attached or are detached from the ER-derived vesicles. Treatment of the ER-derived vesicles with Triton X-100 shows that vicilin peptidohydrolase and vesicle membranes can be physically separated without dissolving the membranes, indicating that the proteinase is soluble within the vesicles. These data support the conclusion that the ER is involved in the transport of ribonuclease and proteinase to the protein bodies.     

Sequence heterogeneity in kinetoplast DNA: Reassociation kinetics

Melting and reannealing of purified kinetoplast DNA (kDNA) from Crithidia fasciculata, Trypanosoma mega, and T. brucei have been studied with an automated optical system. The slow reassociation rate of trypanosome kDNA is due neither to the formation of hyperpolymers nor to mispairing of bases and certainly reflects extensive sequence heterogeneity. Simulation of the reassociation kinetics indicates that the kDNA comprises essentially two kinetic components: a fast renaturing component which might be a common sequence present in all the minicircles and a slow renaturing component which is responsible for minicircle heterogeneity. The rapidly renaturing component is more abundant in Crithidia than in trypanosomes.     

Rapid separation and quantitation of 3',5'-cyclic nucleotides and 5'-nucleotides in phosphodiesterase reaction mixtures using high-performance liquid chromatography

A simple, rapid high-performance liquid-chromatography system for the fractionation and direct quantitation of substrates and products in crude phosphodiesterase reaction mixtures is described. Phosphate buffers and a pellicular anion exchange resin are used at ambient temperature. The method is sensitive, measuring picomoles of products with ultraviolet detection and femtomoles with isotopic measurement, and offers several advantages over the more popular batch sorption and manual methods for measuring phosphodiesterase activity. The time required for analysis, less than 8 min for single substrate reaction mixtures, is a fraction of that required with other chromatographic systems, and precision is +/- 5%. Results of studies with an activatable form of phosphodiesterase demonstrate the accuracy, precision and utility of the procedure for biochemical analyses.     

Zinc mediated effects on leaf CO2 diffusion conductances and net photosynthesis in Phaseolus vulgaris L.

Supra-optimal levels of zinc in primary leaves of Phaseolus vulgaris increased the CO₂ compensation point and inhibited net photosynthesis. Leaf morphology was modified: mesophyll intercellular area, stomatal slit length and interstomatal distance were reduced, but stomatal density increased. Internal and stomatal conductances to CO₂ diffusion decreased. These changes are discussed in relation to the observed effects on leaf gas exchange and to the previously reported inhibition of different photosynthetic and photorespiratory enzymes.     

Relations between substrate feeding pattern and development of filamentous bacteria in activated sludge processes:

Summary Laboratory scale activated sludge systems were operated under regimes of continuous or intermittent feeding of substrate. In a previous paper it was shown that continuously operated systems resulted in the development of filamentous bacteria and bulking sludges. Intermittently fed sludges resulted in good settling. These results are now confirmed when substrates other than glucose are present in the influent, such as nutrient broth, acetate and starch. With casein deflocculation occurred. For intermittent systems the substrate removal rates were higher than for continuous systems. Based on the results a theory is presented to account for the growth of filamentous bacteria (and bulking) in continuous systems (completely mixed systems). This theory assumes that in intermittently fed systems (plug flow systems) floc forming bacteria become dominant as a result of higher substrate uptake rates and the possibility to survive a starvation phase by thriving on accumulated intracellular metabolites.     

Specific inactivation of lysosomal glycosidases in living fibroblasts by the corresponding glycosylmethyl-p-nitrophenyltriazenes.

Glicentin (a highly purified 100-amino acid peptide with glucagon-like immunoreactivity from porcine gut) was subjected to limited digestion with trypsin and carboxypeptidase B, and the resulting peptides were studied by gel filtration and region-specific glucagon radioimmunoassays. Similar digests of glucagon and purified fragments of glucagon were studied in parallel. Glicentin gave rise to peptides that corresponded closely to the 1-17 and 19-29 fragments of glucagon. Also, 125I-labelled glicentin and 125I-labelled glucagon gave rise to identical fragments after trypsin treatment. On the basis of this and other evidence [Jacobsen, Demandt, Moody & Sundby (1977) Biochim. Biophys. Acta 493, 452-459] it is concluded that glicentin contains the entire glucagon sequence at residues number 64-92 and thus fulfills one of the requirements for being a 'proglucagon'.     

Carcinogen aflatoxin B1 is located preferentially in internucleosomal deoxyribonucleic acid following exposure in vivo in rainbow trout

The purpose of this work was to investigate the distribution in chromatin of deoxyribonucleic acid (DNA) adducts of aflatoxin B1, following exposure in vivo. Rainbow trout were injected intraperitoneally with radiolabeled aflatoxin B1, a potent procarcinogen known to readily induced hepatocellular carcinomas in these fish. After maximum incorporation, liver nuclei were prepared and digested with micrococcal nuclease. Mono-, di-, and trinucleosomal fractions were purified from several stages of nuclease digestion, and the lengths and specific activities of their DNA were determined. The results indicate that aflatoxin B1 is approximately 5 times as likely on a per nucleotide basis to localize on internucleosomal (linker) DNA as on nucleosomal core DNA in this system.