DNP-INT

Electron Transport Between Photosystem I and Photosystem II Inhibitors. The interaction between PSI and PSII reaction centers (Fig. 1) depends on the thermodynamically favored transfer of electrons from low redox potential carriers to carriers of higher redox potential. This process serves to communicate reducing equivalents between the two photosystem complexes. Photosynthetic and respiratory membranes of both eukaryotes and prokaryotes contain stmctures that serve to oxidize low potential quinols while reducing high potential metaHoproteins (40). In plant thylakoid membranes, this complex is usually referred to as the cytochrome b /f complex, or plastoquinolplastocyanin oxidoreductase, which oxidizes plastoquinol reduced in PSII and reduces plastocyanin oxidized in PSI (25,41). Some diphenyl ethers, eg, 2,4-dinitrophenyl 2 -iodo-3 -methyl-4 -nitro-6 -isopropylphenyl ether [69311-70-2] (DNP-INT), and the quinone analogues,... 

Fig. 6 (B) shows a recording of neutral-red absorbance changes in chloroplasts, with the initial population present in the Si-state, in response to a flash train. The chloroplasts also contained the inhibitor DNP-INT (dinitrorphenolether of iodonitrotoluol) to suppress plastohydroquinone oxidation and the attendant proton release. The recording is obtained by averaging the responses of 100 fresh chloroplast samples. With a 200-/ -per-channel time resolution, the rapid rise is only partially resolved. A plot of the relative yield ofthe released protons in Fig. 6 (B) actually shows a proton-release pattern of [0,1,2,1 ] for the transition sequence [Si->S2->S3->(S4)->So->S,]. Since the initial, dark-adapted state was 100% in the S, state, the proton release pattern in the [So->S,->S2->S3->(S4)->So] transition is thus [1,0,1,2].

This reaction was inhibited by using DNP-INT (a gift of Prof. Trebst), an inhibitor of the Z-site of the b6-F. 

Fig.l. Absorption changes of NR at 572nm (an increase in absorption corresponds to acidification). Spinach thylakoids in the presence of BSA (10 mg/ml), pH 7.5, and 20 yM DNP-INT. Left time-course during a series of flashes (600 ms apart), with sampling at 5, 30, 100-500 ms after each flash. Right a plot of the change caused by each flash at 5 ms (circles), 30 ms (triangles) and 200 ms (squares). 

Fig.l Relative yield of proton release into the lumen as function of flash number, derived from measurements of absorption changes of neutral red at 548 nm. Heasured in presence of 5pH DNP-INT, 10 pH DMQ and 2.6 mg/ml BSA. Under these conditions absorption changes of neutral red was only due to proton release from water oxidation. 

It is concluded that it is predominantly only one heme, heme that is reduced by a light flash, and that interheme electron transfer was not observed under the conditions of these experiments. The quinone binding site that serves for reduction of heme by PQ- can also serve in the green dga Chlorella as the site for DNP-INT-sensitive oxidation of pre-reduced cytochrome be (9). The use of this pathway for the oxidation of heme bp would be consistent with the conclusion that the electrogenic step under reducing conditions does not arise from interheme transfer (10). One should note that there has been a report of interheme electron transfer in vitro in the isolated cytochrome b6-/complex (11). 

Neither photosystem I nor II were implicated in desaturation, as light is not required. Supporting this concept are the observations that neither DCMU nor atrazine (which prevent reduction of PQ at the Qb site of PS II, fig. 4) inhibited desaturation (fig. 2). Because PQ is involved in "reverse" electron transport, the effects of inhibitors of its function were determined. DBMBIB, UHDBT, BPA, stigmatellin (STG), and DNP-INT interfere with oxidation of reduced PQ by blocking the Qz site (fig. 4), and in fact the first three were able to Inhibit desaturation (fig. 3 and table 5). Both DNP-INT and STG may bind at an overlapping but more... 

Taken together, these results suggest that reduced PQ may be involved in desaturation. However, all of these inhibitors except DNP-INT and STG also act as electron acceptors for FNR (fig 4), and thus could inhibit elsewhere in desaturation by withdrawing electrons from a similar type of flavoproteln. 

Malkin, R (1986) Interaction of stigmatellin and DNP-INT with the Rieske iron-sulfur center of the chloroplast cytochrome bfi-f complex. FEBS Lett 208 317 - 320... 

The rate of cycling cind the redox poise of components in the steady state was dependent on the relative amounts of quinone and cytochrome c (plastocyanin) added and on the number of electrons present in the system. The pH was also critical a pH too high caused significant non-enzymatic electron transfers whereas a pH too low decreased be complex activities. Optimal pH values were around 7.2 for RCbc vesicles and around 6.5 for PSbf vesicles. Under appropriate conditions, redox changes of cytochrome b could also be observed. Antimycin A inhibited cytochrome b reoxidation and myxothiazol inhibited donation into the be complex in the RCbc system, but both were without effect of the PSbf system. Instead 2-n-heptyl-4-hydroxyquinoline-N-oxide could be used to inhibit cytochrome b reoxidation and DNP-INT could be used to inhibit donation into the bf complex. Figure 1 illustrates some typical results. 

Singer SJ (1964) On the heterogeneity of anti-hapten antibodies. Immunochemistry 1 15-20 Soeberg B, Sumerska T, Binns RM, Balfour BM (1978 a) Contact sensitivity in the pig. II. Induction by intralymphatic infusion of DNP-conjugated cells. Int Arch Allergy Appl Immunol 57 114-125... 

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