Evidence by NMR for mobility of the cytochrome domain within flavocytochrome b2

According to a model proposed by Gervais, M, Groudinsky, O., Risler, Y. and Labeyrie, F. ((1977) Biochem. Biophys. Res. Commun. 77, 1543-1551) flavocytochrome b2 is composed of a central flavodehydrogenase entity of 4 X 45 kDa to which are attached four cytochrome b2 globules of approx. 11 kDa that are released after proteolysis of the connective loops. A possible inherent mobility of the latter with functional significance was suspected. Proton NMR spectra at 400 MHz of the isolated and of the flavodehydrogenase-bound ferricytochrome b2 units have been compared. In the ranges downfield of +12 ppm and upfield from -4 ppm, where hyperfine-shifted heme proton resonances reside, the chemical shifts are identical for the two forms, but the linewidths are markedly broader for flavocytochrome b2. The linewidths of three heme resonances, a methyl at +19 ppm, two single protons at -6 and -8 ppm (most probably from one vinyl) and an unassigned line at -2.4 ppm, all increase by a factor of about 4. Since, in the present case, linewidths are controlled mainly by proton/proton dipolar relaxations which are caused by molecular tumbling, a change in linewidths of about 15 would be expected if the cytochrome b2 globule had no free motion relative to the flavodehydrogenase domain. The present results thus support the previous hypothesis that such a relative mobility, of unknown correlation time and amplitude, actually exists.     

Modification of redox equilibria between heme and flavin within yeast flavocytochrome b2 (l-lactate cytochrome c reductase) upon binding of pyruvate,the reaction product

Direct determinations of the concentration of semiquinone spin in redox equilibrium with the cytochrome b₂ moiety were carried out at room temperature in the presence of added pyruvate or in its absence. Results show that redox potentials of the one-electron couples of the prosthetic flavin are markedly affected by binding of pyruvate the reaction product in the oxidation of l-lactate. The proportion of flavin semiquinone nearly reaches then 100 per cent.     

Modifications of redox equilibria with semiquinone stabilization upon pyruvate binding to L-lactate cytochrome c oxidoreductase (flavocytochrome b2)

Spectral redox titrations of flavin and cytochrome b₂ moieties of flavocytochrome b₂ were achieved in the absence and in the presence of pyruvate under equilibrium conditions at 18° C; direct measurements of spin flavosemiquinone proportions have been carried out by EPR determinations at the same temperature. Our results show that the equilibria involving flavin are largely affected by the presence of pyruvate; the semiquinone proportion markedly increases almost till unit near half-reduction of cytochrome b₂; at 10 mM pyruvate, the dismutation constant, $\text{K}{}_{\mathrm{dism}}\text{=}\text{(}\text{F}{}_{\mathrm{s}}\text{)}{}^2\text{(}\text{F}{}_{\mathrm{o}}\text{)}{}^1\text{(}\text{F}{}_{\mathrm{r}}\text{)}$ increases by a factor ≥ 10.     

Remarques sur la mise au point d'un élevage semi-industriel deMacrocentrus ancylivorus Roh

Summary Inspired by the breeding technique ofFinney, Flanders andSmith, the one which we have elaborated can be carried out by a reduced personnel, thanks to an increase of the output. The adults of theGnorimoschema operculella that are fed lay 300 eggs, most of them within 4 days. The limited keeping of the eggs and adults of the moth is possible at low temperatures and depends on the deficit of saturation of air moisture. The limited keeping of the cocoons ofM. ancylivorus containing larvae in their last stage can be assured at low temperatures. The notion of breeding index based on the variations of the rate of multiplication of the parasite for given relative densities makes it possible to check the breeding conditions. The average rate of multiplication which has been obtained is 76. The breeding technique must allow for the absolute necessity of preventing the extension of possible epidemics this implies a choice of desinfectable materials, the rotation of breeds, the separation of stages, the division of breeds in separable groups, the isolation of soiled implements, the periodical disinfection of the premises and materials. The technique we make use of enables a single man to obtain an output of 280 000 cocoons ofM. ancylivorus per month.

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Resumen Inspirada en la técnica de cría deFinney, Flanders y Smith, la que hemos elaborado puede ser aplicada por un personal limitado merced a un aumento de los rendimientos. Los adultos deGnorimoschema operculella alimentados, ponen 300 huevos, la mayor parte en 4 días. La conservación limitada de los huevos y de los adultos es posible con bajas temperaturas y depende del déficit de saturación en vapor de agua. La conservación limitada de los capullos deM. Ancylivorus que contienen larvas en su último estado se puede obtener conbajas temperaturas. La noción de índice de cría basada en las variaciones del coeficiente de multiplicación del parasito para densidades relativas dadas permite la verificación de las condiciones de cría. Ha sido de 76 el coeficiente medio de multiplicación. En la técnica de cría, se debe tomar en consideración la necesidad absoluta de precaver la extensión de eventuales epidemias. Esto supone la elección de materiales desinfectables la permutación de las crías, la separación de los estadios, el fraccionamiento de las crías en grupos, aislables, el aislamiento del material ensuciado, la desinfección periódica de los locales y materiales. La técnica utilizada permite la producción de 280 000 capullos deM. ancylivorus al mes por solo un individuo.