Applicability of the induced-fit model to glyceraldehyde-3-phosphate dehydrogenase from sturgeon muscle. Study of the binding of oxidized nicotinamide adenine dinucleotide and nicotinamide 8-bromoadenine dinucleotide

A new method of calculation, based on a direct fitting of the protein fluorescence intensity observed upon coenzyme binding (H.-P. Lutz, unpublished results), is used to study the negative cooperative behavior of glyceraldehyde-3-phosphate dehydrogenase from sturgeon muscle. The calculation procedure simultaneously elaborates data obtained for four different protein concentrations, and it is able to compare different models by computing the minimal and critical sum of squares. Using this approach, it is shown that the induced-fit model [Koshland, D. E., Jr., Nemethy, G., & Filmer, D. (1966) Biochemistry 5,365] and the dimer of dimer model [Malhotra, O. P., & Bernhard, S. A. (1968) J. Biol. Chem. 243, 1243-1252] can both be applied for explaining the negative cooperativity observed upon coenzyme binding to sturgeon glyceraldehyde-3-phosphate dehydrogenase. In addition to the progressive modification of the binding affinity during ligand binding, different maximal fluorescence quenchings for the binding steps must be postulated; and furthermore, the binding capability decreases by decreasing the protein concentration. The fact that the induced-fit model can also be applied is rather in contradiction with the view generally accepted of a dimer of dimer structure of sturgeon glyceraldehyde-3-phosphate dehydrogenase. By use of the same approach, nicotinamide 8-bromoadenine dinucleotide is shown to bind to glyceraldehyde-3-phosphate dehydrogenase from sturgeon in a negative cooperative manner.     

Comparison of the effect of temperature on kidney cortex mitochondria from rabbit,dog, pig,and human: Arrhenius plots of ADP-stimulated respiration

The effect of temperature on the rate of ADP-stimulated respiration of mitochondria from dog, rabbit, pig, and human kidney cortex mitochondria was plotted according to the Arrhenius relationship. The temperature at which the plot demonstrated a break was at 15 °C for mitochondria from dog, pig, and human kidneys. The discontinuity occurred at 10 °C or less for mitochondria from rabbit kidneys. This difference suggests that mitochondria from rabbit kidneys undergo a lipid-phase transition at lower temperatures than for other species commonly used in experimental renal preservation. The implications of this difference suggest caution in using results obtained with rabbit kidneys for comparison to results obtained from hypothermic renal preservation of other species kidneys. Apparent fluidization of dog kidney mitochondrial membranes with adamantine abolished the discontinuity in the Arrhenius plot.     

The partial purification and characterization of cytosol alcohol dehydrogenase from Astasia

1. The cytosol alcohol dehydrogenase (alcohol-NAD oxidoreductase, EC 1.1.1.1) of Astasia longa was partially purified and characterized from cells grown in the presence of air+CO(2) (95:5) or of O(2)+CO(2) (95:5). 2. Under both these growth conditions, the cells contained a fraction, ADHII, which was characterized by its electrophoretic properties, by a high degree of resistance to heat inactivation, by a sharp pH optimum at 8.2 and by its kinetic properties. The estimated molecular weight of this fraction was approx. 150000, which is similar to that of yeast alcohol dehydrogenase. 3. Cells grown in air+CO(2) (95:5) contain another fraction, ADHI, which can be further separated into two subfractions by polyacrylamide-gel electrophoresis and by DEAE-cellulose chromatography. This was termed fraction ;ADHI-air'. 4. In addition to fraction ADHII, cells grown in the presence of O(2) have a twofold increase in fraction ADHI-air activity as well as two new fractions that could not be demonstrated in air-grown cells. These new fractions which we have called fraction ;ADHI-O(2)', account for about 10% of the total activity. 5. The ADHI fractions (air) and (O(2)) have similar broad pH-activity curves and similar kinetic properties, both having a lower K(m) for ethanol and NAD than fraction ADHII. However, they differ from each other with respect to their activity with various substrates. The estimated molecular weight of these two ADHI fractions and their chromatographic behaviour on hydroxyapatite and on DEAE-cellulose also distinguish them.     

Hg ++ - a DCMU independent electron acceptor of photosystem II

Mercuric chloride functions as a direct electron acceptor from the quencher of fluorescence in Photosystem II. The photoreduction of ferricyanide, dichlorophenol-indophenol or methyl viologen is inhibited by mercuric ion while oxygen evolution is uneffected. Mercuric chloride supported oxygen evolution (mercury Hill reaction) is not prevented by DCMU or other similar electron transport inhibitors.     

Über die Herkunft „synaptischer“ Bläschen in neurosekretorischen Axonen

Zusammenfassung In der Neurohypophyse fetaler und neugeborener Ratten entsteht die Mehrzahl der synaptischen Bläschen aus Erweiterungen der Neurotubuli. Ferner können pinocytotische Bläschen als synaptische Vesikel imponieren. Die Bläschenbildung aus Membranen von Elementargranula (vgl. Herlant, 1967) tritt dagegen in den Hintergrund. Ein Auftreten von Vesikeln im Innern von Elementargranula wurde nicht beobachtet.

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The origin of synaptic vesicles in neurosecretory axons

Summary In the neurohypophysis of fetal and newborn rats the majority of synaptic vesicles originate from dilatations of neurotubuli. Moreover, pinocytotic invaginations give rise to synaptic vesicles. Evaginations of elementary granule membranes, as described by Herlant (1967), are seldom to be found and do not seem to play an important role in the formation of synaptic vesicles. The occurrence of vesicles within elementary granules was not observed.

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Mit Unterstützung durch die Deutsche Forschungsgemeinschaft.     

Der cis-trans-Positionseffekt in Heterokaryen morphologischer Mutanten von Podospora anserina

Zusammenfassung Der cis-trans-Positionseffekt in Heterokaryen der nicht gekoppelten morphologischen Mutanten z und oct-1 von Podospora anserina wird durch quantitative Kernunterschiede vorgetäuscht.Im cis-Fall führt die Wuchsverminderung von Hyphen mit überwiegendem Gehalt an Doppelmutantenkernen nach Mikrosektorenbildung im Mycel zu einer Selektion der Hyphen mit überwiegendem Wildkerngehalt. Die Doppelmutantenkerne sind nach 6 cm Wuchsstrecke aus dem Mycel verschwunden. Dieses weist daher Wildphänotyp auf.Im trans-Fall wird eine Kernentmischung durch Anastomosenbildung kompensiert, da die entestehenden Kleinsektoren gleich schnell wachsen. Das Mycel bringt daher die oct-1- und z-Merkmale in einem intermediären Phänotyp zum Ausdruck.

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The cis-trans position effect in heterocaryons of morphological mutants of Podospora anserina

Summary The cis-trans position effect of the two non-linked morphological mutants z and oct-1 in heterocaryons of Podospora anserina is caused by quantitative differences of the two types of nuclei.In the cis configuration, there is a growth diminuition of hyphae containing predominantly nuclei of the double mutant type which leads to a selection of hyphae containing mainly the wildtype nuclei. This is due to the formation of microsectors. After 6 cm of mycelial growth the double mutant nuclei are no longer distinguishable; the mycelium adopts the wild phenotype.In the trans configuration, the microsectors of both types of mutant nuclei exhibit the same growth rate; the separation of nuclei is prevented as a result of the formation of hyphal anastomoses. The mycelium shows then an intermediate phenotype with both mutant characteristics.

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Mit Unterstützung der Deutschen Forschungsgemeinschaft.     

81. Jahresversammlung (1968) zu Innsbruck

Ohne Zusammenfassung     

Absence of Free Aldehydes in Fresh Tissues, as Shown with a Neutralized Schiff Reagent

A neutralized Schiff's reagent (pH 6.7) was prepared and used to investigate the role of the acidic nature of the routine Schiff's reagent (pH 2.6) in the plasmal reaction. The neutralized reagent was satisfactory as an aldehyde reagent in the nucleal reaction on gut and, although giving a less intense reaction than the routine reagent in the PAS reaction on the gut and plasmal reaction on the aorta, was satisfactory here in respect to localization and thus to aldehyde specificity. Control sections for the plasmal reaction of unfixed nerve and aorta gave positive results when placed in the routine Schiff's, this increasing with time left in the reagent. Similar control sections in the neutralized Schiff's reagent remained consistently negative even though left in this reagent for 0.5 hr. The positive reaction of such control sections is apparently due to acid hydrolysis of labile plasmalogens by the routine Schiff's reagent in myelin and elastin and not to the presence of “free” aldehydes in these tissue elements     

Endocytosis in the ureteric duct epithelium of the hagfish (Myxine glutinosa L.)

Summary Solutions containing ferritin or thorotrast particles were microperfused through the ureteric duct of the hagfish. The markers were taken up by the epithelial cells by way of endocytosis and were transported in bulk in apical vesicles. Newly formed apical vesicles containing marker showed bristle coating on the cytoplasmic side of their limiting membrane. This coating appeared to be lost during the movement of vesicles deeper into the cytoplasm.The findings indicate that the epithelial cells in the ureteric duct have capablity for extensive bulk uptake of macromolecules from the luminal fluid. The mechanisms involved in absorption appear to be similar to those in proximal convoluted tubules of mammals.The apical dense tubules observed with some fixation techniques appear to represent collapsed endocytotic vesicles.The authors are indebted to Finn Walwig, Cand. real., Marine Biology Station, University of Oslo, Dröbak, Norway for kindly supplying the hagfishes used in this study. The technical assistance of Miss Signe Fjeldsenden and Miss Britt-Marie Pettersson is gratefully acknowledged.This work has been supported by grants from the Karolinska Institutet Medical School, Stockholm, Sweden (Therese och Johan Anderssons Minne).