Abstract
The enzyme NADPH:protochlorophyllide oxidoreductase (POR) catalyses the reduction of protochlorophyllide into chlorophyllide, a precursor of chlorophyll a in photosynthetic organisms. The enzyme binds the substrate and the cofactor in the dark and catalysis is initiated by the absorption of light by the substrate. We have carried out spectroscopic measurements with ultrafast time resolution under single pulse conditions, which reveal a biphasic formation of the first catalytic intermediate, I675* with average rates of (3.7 ± 0.7 ps) -1 and (177 ± 78 ps) -1, as obtained from a systematic analysis of 15 datasets. Measurements in the mid-IR absorption spectral region show that I675* is associated with a decrease of the PChlide CO keto oscillator strength. The spectroscopic changes in the visible and mid-IR regions are specific for the enzyme reaction as they do not occur in the photoexcited substrate alone. In deuterated samples, the rates of I675* formation are reduced by a factor of 1.5-2 compared to protonated samples, suggesting the involvement of a proton movement in this reaction step. The quantum yield of this step is determined to be 0.64 ± 0.11, and the quantum yield of the final reaction product formed on a later time scale, chlorophyllide, is 0.26 ± 0.06. Several possible interpretations of these data are discussed. © 2011 The Owner Societies.
Original language | English |
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Pages (from-to) | 616-625 |
Number of pages | 9 |
Journal | Physical Chemistry Chemical Physics |
Volume | 14 |
Issue number | 2 |
DOIs | |
Publication status | Published - 14 Jan 2012 |