Study of the Electron- and Proton Transport in Inside-Out Thylakoids

Abstract

Inside-out thylakoids are an attractive system for the investigation of the functional sidedness of the thylakoid membrane, especially for the study of protolytic reactions coupled with wateroxidation which occur at the inner side of normal thylakoids. The functional arrangement of the electron transport chain of these particles is not yet clarified. Therefore, comparative pulse spectroscopic investigations of the following components of the chain in normal and inside-out thylakoids have been performed: P 680, X 320, PQ (Plastoquinone) and P 700. It was found that the inside-out thylakoids are slightly enriched in PS II (P680/P700 ≳ 1) and that the connection between the PQ-pool and PS I (P 700) is interrupted. Accordingly, these particles mainly behave as PS II — vesicles. The effect of trypsin on inside-out thylakoids was studied in a similar way as was done before on normal chloroplasts (1). In contrast to normal chloroplasts the reoxidation kinetics of X 320⁻ in inside-out vesicles is not affected by trypsination. This result provides a further proof that the membrane is turned inside-out (former proofs 2, 3, 4). Surprisingly, the watersplitting enzyme system Y seems to be rather stable against trypsination which is shown by oxygen and proton measurements. In inside-out particles the overall half rise time of the proton release due to wateroxidation is found to be 1 ms which is in accordance with measurements on normal chloroplasts. Preliminary results on the oscillatory pattern of this proton release will be discussed.