The Relationship of Internal Conductance and Membrane Capacity to Mitochondrial Volume

A study was made of the effect of mitochondrial size on the electrical properties of the membrane and the internal conductivity of mitochondria. The dielectric constant and electrical conductivity of suspensions of guinea pig heart mitochondria were examined in the frequency range 5 x 10⁵ to 2.5 x 10⁸ C.P.S. Membrane capacity was calculated to be 1.1 to 1.3 µf./cm.² and was virtually the same in mitochondria whose surface area was made to vary by a factor of 4 by osmotic means. This finding suggested that some mechanism must exist for the transfer of mitochondrial material into membrane structure during fluctuations in mitochondrial size. The electrical capacity of the membrane was unaffected by a 33-fold change in potassium chloride concentration. The internal conductance of swollen mitochondria was 2 to 3 times lower than that of the external medium. In shrunken mitochondria the internal conductance was virtually independent of the conductivity of the external medium. These results were discussed in relation to current concepts of mitochondrial structure.     

Schmelzverhalten und kalorische Eigenschaften einer Mischung von Rinderserumalbumin,Kochsalz und Wasser

Zusammenfassung Es wurde die mittlere EnthalpiedifferenzH _m von Rinderserumalbumin (RSA)-Salz-Wassergemischen im Temperaturbereich von –47 °C bis 0°C bestimmt.H _m enthält neben der Energie zur Erwärmung auch noch die Schmelzwärme der Mischung. Aus dem Verlauf vonH _m als Funktion des Lösungsmittels (0,5 molale NaCl-Lösung) ergibt sich für den Anteil des kalorisch gebundenen Lösungsmittels ein Wert von 0,54 g Lösungsmittel pro g RSA. Dieser Anteil ist unabhängig von der mittleren Ladung des RSA-Moleküls.Aus einer Analyse der Meßergebnisse nach der Theorie der Mischungen folgt, daß die Partialenthalpie des RSA in verdünnten Mischungen negativ ist. Die Partialenthalpie des Lösungsmittels ist dagegen in diesen verdünnten Gemischen unverändert gleich der Enthalpiedifferenz des Lösungsmittels selbst. Die Messungen wurden mit Hilfe eines Differential-Scanning-Kalorimeters der Fa. Perkin-Elmer durchgeführt.

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Melting behaviour and caloric properties of a mixture of bovine serum albumin, sodium chloride and water

Summary In the temperature range from –47 °C to 0 °C the enthalpy differencesH _m of bovine serum albumin (BSA)-salt-water mixtures were measured.H _m includes the energy to increase the temperatures as well as the melting heat of the mixture. The plot ofH _m as a function of the solvent content (0,5 molal NaCl solution) gives 0,54 g solvent per g dry BSA for the caloric bound solvent. This value is independent of the average charge of the BSA molecule.The theory of mixtures applied to this measurements shows that the partial enthalpy difference of BSA in dilute mixtures is negative while that of the solvent in this mixtures is equal to the enthalpy difference of the pure solvent.The enthalpy differences were measured with a Differential-Scanning-Calorimeter (Perkin-Elmer).

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Herrn Prof. Dr. Dr. h. c. mult. Boris Rajewsky zum 80. Geburtstag gewidmet.     

Molecular dynamics in protein-single stranded DNA complexes. Two distinct nucleoside mobilities in poly(deoxythymidylic acid)-gene 5 protein complexes

A set of differently spin labeled (dT)n is used to evaluate thymidine dynamics and some of the structural features in a (dT)n-gene 5 protein complex. ESR evidence is presented that only one of the four thymidine residues bound in the DNA binding channel shows strong immobilization, whereas the other three display significant mobility of the order of nanoseconds. It is hypothesized that the accessability of such mobile bases could be critical to the recognition of the (dT)n-gene 5 protein complex in auxiliary interactions with other proteins and competitive DNAs.     

Elektrolyttransport in Ehrlich-Aszitestumorzellen nach Röntgenbestrahlung

Zusammenfassung In der vorliegenden Arbeit wurde die Röntgenstrahleneinwirkung auf den Kaliumtransport bei Ehrlich-Aszitestumorzellen untersucht. Die Zellen wurden im dichtgepackten und anaeroben Zustand bestrahlt. Es waren Strahlendosen von einigen 100 krad erforderlich, um den Kaliumtransportmechanismus zu beeinträchtigen. Nach einer Strahlendosis von etwa 350 krad betrug der Kaliumgehalt nur noch 50% der Kontrolle. DerEfflux-koefnzientk _ex =0,04 min^–1 nahm um 50% zu, während derInfluxkoeffizientk _in =1,04 min^–1 nur um 20% abnahm. Nach Strahlendosen von mehr als 500 krad konnten die Zellen kein dynamisches Gleichgewicht mehr aufrechterhalten.

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The effect of x-rays on the electrolyt-transport in Ehrlich- Ascites tumor cellsIrradiation of tightly packed cells

Summary This paper deals with the radiosensitivity of the potassium transport in Ehrlich -Ascites tumor cells. During the irradiation the cells were packed tightly and kept under anaerobic conditions. To reduce the potassium content very large doses were required. After a dose of 350 krad the potassium content decreased to 50% of the control value. The efflux coefficientk _ex =0,04 min^–1 increased by about 50%, and the influx coefficientk _in =1,04 min^–1 decreased by about 20%. After doses higher than 500 krad the cells were unable to maintain a dynamic equilibrium.

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Herrn Professor Boris Rajewsky zum 75. Geburtstag gewidmet.

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Der vortiegenden Ver6ffentlichung wurde die Dissertation yon G.Itasl [10] zugrunde gelegt. Die experimentel]en Arbeiten wurden in den Jahren 1963 bis 1967 im Max-Planck-Institut für Biophysik in Frankfurt am Main unter seinem Direktor Prof. Dr. B.Rajewsky durohgefiihrt.     

Das Membranpotential von Ehrlich-Aszitestumorzellen

Zusammenfassung Mit Hilfe von Mikroelektroden wurde das Membranpotential von Ehrlich-Aszitestumorzellen in Krebs-Ringer-Phosphatlösung (KRP) bei 25 °C bestimmt. Die gemessene Potentialdifferenz der gesamten galvanischen Zelle enthält außer dem Membranpotential selbst noch zwei spezielle Diffusionspotentiale an den Mikrokapillaren, welche zur Ermittlung des Membranpotentials bekannt sein müssen. Das Diffusionspotential an der Bezugskapillare ist klein und konnte mit genügender Genauigkeit berechnet werden. Die Spitzenpotentiale der Mikrokapillaren (Spitzendurchmesser etwa 0,5 m) sowie deren Änderung beim Übergang von KRP in das Zytoplasma wurden experimentell bestimmt. Diese Änderung beträgt 37% des Spitzenpotentials selbst, und zwar für Spitzenpotentiale bis zu 70 mV. Mit diesen Korrekturen wurde das Membranpotential zu –11,5 mV ±5% bestimmt. Die angegebene Methode der Korrektur erlaubt es, das Membranpotential mit guter Genauigkeit auch dann zu messen, wenn Mikrokapillaren mit Spitzenpotentialen bis zum fünffachen Wert des Membranpotentials verwendet wurden. Somit ist die Messung von Membranpotentialen sehr kleiner Zellen mit Hilfe von extrem spitzen Mikrokapillaren möglich, deren Spitzen gewöhnlich größere Spitzenpotentiale zeigen.

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Summary The membrane potential of Ehrlich-mouse ascites tumor cells in Krebs-Ringer-Phosphate solution (KRP) at 25 °C have been measured to be –11.5 mV ±5%. This value was obtained after correcting the measured electromotive force of the total galvanic cell including two diffusion potentials and the membrane potential for the tip potential of the microelectrode used with a tip of 0.5 m diameter, and for the change in tip potential from the value in KRP to the value in the cyctoplasma. Experimentally it could be shown that this change in tip potential is 0.37 times the value of the tip potential itself for tip potentials as high as 70 mV. The small value of the diffusion potential of the reference electrode have been calculated with sufficient accuracy. By this method it was possible to measure the membrane potential with good accuracy with microelectrodes having tip potentials five times as large as the membrane potential itself. Hence it should be possible to measure the membrane potential of very small cells with correspondingly fine microelectrodes having usually higher tip potentials.

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Der vorliegenden Veröffentlichung wurde die Diplomarbeit von J.Bernhardt [2] zugrunde gelegt. Sie wurde von 1964 bis 1966 im Max-Planck-Institut für Biophysik in Frankfurt am Main unter seinem Direktor Prof. Dr. B.Rajewsky durchgeführt.     

Electrical Properties of Mitochondrial Membranes

The electrical capacity of the membrane of rat liver mitochondria is 0.5 to 0.6 µ./cm². This membrane capacity is obtained from the analysis of the frequency dependence of the admittance of a suspension of swollen mitochondria. In potassium chloride media the mitochondrial membrane capacity does not depend on the ion concentration. The internal conductance of the mitochondria was approximately one-half that of the external medium; the same applies if the mitochondria are equilibrated in a medium with a 10-fold difference in potassium chloride concentration. Hence the swollen mitochondria investigated here appear to be able to adjust their internal ion concentration in proportion with that of the external phase. The similarity of the membrane capacity of isolated mitochondria with the range of values known for other membranes suggests a common molecular structure. The analysis of experimental data suggests an anisotropic electrical behavior of the interior of mitochondria. This anisotropy is readily explained by the existence of internal membranes.     

Immunofluorescent patterns of spectrin in lymphocyte cell lines

Spectrin, a membrane-associated cytoskeletal protein, has been observed in all of 45 lymphoid and myeloid cell lines examined. For these experiments, formalin-fixed cells from randomly selected lines propagated by using conventional tissue culture procedures were examined by immunofluorescence, using an antibody directed against chicken erythrocyte alpha-spectrin. Two distinct immunofluorescent patterns of spectrin distribution were identified. In most lines examined (16 mouse and 18 human lymphoid or myeloid lines), spectrin was symmetrically distributed near the submembranous region of the plasma membrane. In the remainder of the cell lines examined, a second pattern was observed; in these cultures, the cells contain a polar submembranous aggregate of spectrin with little staining at the rest of the plasma membrane. Long-term T lymphocyte cell lines in which greater than 60% of the cells expressed a polar submembranous aggregate of spectrin (PSA-S) include mouse cell lines EL-4, LBRM-33, CT-6X, NIXT, 22CM-37, and 7ON-2 and human lines JM and PEER. Other established cultures in which PSA-S were observed included the human macrophage-like line U-937 and gibbon T cell line MLA-144. Phorbol myristate acetate or mezerin caused a reversible alteration in the distribution of spectrin in these cell lines. These drugs, which increase membrane fluidity, caused a complete but temporary symmetrical redistribution of the spectrin aggregate. Our results indicate that the pattern of spectrin distribution, either aggregated or evenly dispersed, is a stable characteristic (but one that can be altered) in various cell lines, and that because similar variations in pattern have been noted in situ, it is likely that the pattern present in any given cell line reflects a characteristic associated with a particular stage of a cell's maturation. It is anticipated that these cell lines, positive and negative for the expression of natural polarity of spectrin distribution, will provide useful models for future studies to define further the role of spectrin in lymphocyte plasma membrane functions.     

Production and characterization of a monoclonal antibody to the trichothecene mycotoxin diacetoxyscirpenol

A monoclonal antibody was obtained by the fusion of mouse myeloma cells with splenocytes isolated from Balb/c mice, which had been immunized with diacetoxyscirpenol-hemiglutarate (DAS-hemiglutarate) and verrucarol-hemiglutarates covalently bound to ethylenediamine-modified bovine serum albumin. The anti-DAS-antibody that could be induced was of the IgM type with kappa-chains. The titer of the monoclonal anti-DAS-antibody in ascites fluid obtained from mice injected the selected cell line was much higher than those of conventional antisera. An enzyme-linked immunosorbent assay based on the competitive binding principle in which the antibody was applied had a sensitivity of 1 ng DAS per assay. The relative cross-reactivity of the monoclonal antibody in the CI-ELISA with the related trichothecenes such as triacetoxyscirpenol, 15-monoacetoxyscirpenol, diacetylverrucarol, 4-monoacetoxyscirpenol and scirpentriol were found to be 1.8, 0.8, 0.15, 0.02 and less than 0.001, respectively. The trichothecenes verrucarol, T-2 toxin, T-2 tetraol, deoxynivalenol, 3-acetyldeoxynivalenol and trichothecin showed no cross-reactivity.