. Biological structure and function; proceedings. Biochemistry; Cytology. SOLUBILIZATION AND PROPERTIES OF THE DPNH DEHYDROGENASE lOQ of ETP or DPNH oxidase preparations to this inhibitor in the flavoprotein- ferricyanide interaction (cf. discussion of Fig. i) suggest that amytal interrupts electron transport between flavoprotein and the respiratory chain, as is also the case in the choline oxidase system of liver [17], and not between DPNH and flavoprotein. Contrary conclusions in the earlier literature were based on the "cross-over technique" which relies on the measurement of the
. Biological structure and function; proceedings. Biochemistry; Cytology. SOLUBILIZATION AND PROPERTIES OF THE DPNH DEHYDROGENASE lOQ of ETP or DPNH oxidase preparations to this inhibitor in the flavoprotein- ferricyanide interaction (cf. discussion of Fig. i) suggest that amytal interrupts electron transport between flavoprotein and the respiratory chain, as is also the case in the choline oxidase system of liver [17], and not between DPNH and flavoprotein. Contrary conclusions in the earlier literature were based on the "cross-over technique" which relies on the measurement of the oxidation state of the flavoprotein in the 450-465 mfj. region. As will be documented later in this paper, the application of this technique to DPNH dehydrogenase has some major weaknesses: the diff'erence spectrum is atypical of simple flavoproteins; the extinction coefficient of simple flavoproteins at 465 m^u, is not applicable to this enzyme; and it is not even certain that the flavin is fully reduced in normal catalysis. 40 NAJA-NAJA 1:25. (mm) Fig. 6. Progress of solubilization of DPNH dehydrogenase by phospholipase A. Aliquots of a DPXH oxidase preparation [11] were incubated at 30^, pH 7-4, with Naja naja venom as a source of phospholipase. The ratios indicated define the mg. weight of venom employed per mg. protein in the particulate preparation (determined by biuret reaction, using a coefficient of 0-095 for i nng. protein per 3 ml., I cm. light path, 540 m/w). At various times aliquots were rapidly cooled to o , centrifuged at 105 000 x g for 30 min., and the supernatant solution was assayed. Activities are given in arbitrarv units on the ordinate. Turning now to the isolation and characteristics of the dehydrogenase, Fig. 6 shows the progress of solubilization of the enzyme, starting with ETP, at two levels of cobra venom. Compared with brain a-glvcerophos- phate dehydrogenase [9] and choline dehydrogenase from liver [18], the level of phospholipase A (cobra venom)
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