Nuclear functions required for cytochrome c oxidase biogenesis in Saccharomyces cerevisiae. Characterization of mutants in 34 complementation groups

To identify nuclear functions required for cytochrome c oxidase biogenesis in yeast, recessive nuclear mutants that are deficient in cytochrome c oxidase were characterized. In complementation studies, 55 independently isolated mutants were placed into 34 complementation groups. Analysis of the content of cytochrome c oxidase subunits in each mutant permitted the definition of three phenotypic classes. One class contains three complementation groups whose strains carry mutations in the COX4, COX5a, or COX9 genes. These genes encode subunits IV, Va, and VIIa of cytochrome c oxidase, respectively. Mutations in each of these structural genes appear to affect the levels of the other eight subunits, albeit in different ways. A second class contains nuclear mutants that are defective in synthesis of a specific mitochondrial-encoded cytochrome c oxidase subunit (I, II, or III) or in both cytochrome c oxidase subunit I and apocytochrome b. These mutants fall into 17 complementation groups. The third class is represented by mutants in 14 complementation groups. These strains contain near normal amounts of all cytochrome c oxidase subunits examined and therefore are likely to be defective at some step in holoenzyme assembly. The large number of complementation groups represented by the second and third phenotypic classes suggest that both the expression of the structural genes encoding the nine polypeptide subunits of cytochrome c oxidase and the assembly of these subunits into a functional holoenzyme require the products of many nuclear genes.     

Lipid peroxidation in liver, plasma, and erythrocytes of rats chronically treated with ethanol

The effect of ingestion of water containing 20% ethanol for 1-2 months on lipid peroxide levels of liver, plasma, and erythrocyte was investigated in rats. Our results show that elevated plasma lipid peroxide levels and erythrocyte susceptibility to lipid peroxidation may reflect stimulated lipid peroxidation in rat liver following chronic ethanol ingestion.     

Isolation of specific antigens from Angiostrongylus cantonensis. 1. Preparative flatbed isoelectric focusing

Electrophoresis on SDS gel and analytical isoelectric focusing showed that a crude extract of Angiostrongylus cantonensis consisted of at least 40 protein components with molecular weights ranging from 13 000-70 000 and isoelectric points of pI values ranging from 3.7-10.0. Crossed-immunoelectrophoresis with a hyperimmune antiserum to A. cantonensis showed at least 40 different antigenic components in the crude worm extract which were cross-reactive with those of Ascaris suum, Metastrongylus apri and Toxocara canis. Using preparative isoelectric focusing, the somatic worm preparation was divided into 13 equal fractions, of which 3, 4 and 5, with pI values of 3.7, 4.0 and 4.45 respectively, were later shown by immunoelectrophoretic techniques and enzyme-linked immunosorbent assay to contain antigens specific to A. cantonensis.     

Spontaneous Mutations Modifying the Activity of Alcohol Dehydrogenase (Adh) in DROSOPHILA MELANOGASTER

In a marked-inversion balanced lethal system of the second chromosome of Drosophila melanogaster, mutations were accumulated under minimum pressure of natural selection in 1000 individual lines that originated essentially from two individuals. After about 300 generations, the specific activities of alcohol dehydrogenase of 69 randomly selected individual lines were measured with replications using four replicated vials (on 2 days—two replications per day) by observing the reduction of NAD⁺ to NADH at 340 nm. Total soluble protein as the basis of standardization of enzyme activity was measured by the Lowry method for each vial. A control experiment was made immediately after the establishment of 20 individual lines from a single genotype. A significant increase in genetic variance was observed among the mutation-accumulating lines but was not detected in the control experiment. The statistical analysis of the data on the basis of the one-band/one-gene hypothesis suggests that many mutations controlling the activity of alcohol dehydrogenase occurred in regions different from the alcohol dehydrogenase locus itself, mainly in the noncoding DNA. Furthermore, it is suggested that transposon-like elements are related to the induction of these changes in alcohol dehydrogenase specific activities. Additional experimental evidence supporting this conclusion is also given.     

Microbial Production of Lysine and Threonine from Whey Permeate

Extracellular accumulation of lysine and threonine was investigated in modified whey permeate by using Brevibacterium lactofermentum ATCC 21086 and Escherichia coli ATCC 21151. Whey permeate was prepared from whey by membrane ultrafiltration, and lactose was hydrolyzed by treating permeate with HCl or β-galactosidase. The highest amount of lysine (3.3 g/liter) was produced from a mixture of acid-hydrolyzed whey permeate and yeast extract (0.2%). The highest amount of threonine (3.6 g/liter) was produced from a mixture of whey permeate, (NH₄)₂SO₄ (1.4%), yeast extract (0.1%), and Na₂CO₃ (0.3%).     

Expression of zein in long term endosperm cultures of maize

Continuous cultures, established 10 days after pollination from endosperms of inbred A636 Zea mays (L.) were extracted 21 months later with aqueous ethanol. The solubilized proteins were analyzed by poly-acrylamide-sodium dodecyl sulfate gel electrophoresis. Two protein bands co-migrated with zein, the major storage protein of maize. Immunoblotting of the gel followed by incubation of the immobilized proteins with anti-zein IgG provided evidence that the polypeptides were in fact zein. Electron microscopic studies showed that the cultures contained cells with protein bodies as found in developing endosperms. The protein bodies could be isolated from the cultures and were shown to contain zein. We conclude that the long term cultures described here synthesize zein and deposit it in the form of protein bodies of the type found in developing endosperms. Thus, certain endosperm characteristics and the production of tissue-specific proteins are retained in prolonged culture.     

Absorbance-temperature profile of ribosomes from seeds of Pinus lambertiana

The absorbance (260 nm)-temperature profile of ribosomes from seeds of $\text{Pinus lambertiana}$, in the temperature range of about 25 – 60° was observed to be biphasic. When r-proteins in ribosomes were dansylated by 5-dimethylamino-1-naphthalenesulfonyl chloride dispersed on celite (180 molecules of 5-dimethylamino-1-naphthalenesulfonyl chloride per one ribosome particle), each transition midpoint was lowered by 2 – 3° and the sharpness of the transition of each phase was increased. The result of the present work suggests discrete cooperative transitions of the conformation of r-RNAs in ribosomes and its control by r-proteins.