Determination of the Activation Parameters of the Early Steps of Greening in Dark-grown Wheat Leaf Homogenates by High Pressure Fluorescence Spectroscopy
Advances in High Pressure Bioscience and Biotechnology II, 2003
In dark-grown higher plants the chlorophyll biosynthetic pathway stops at protochlorophyllide bec... more In dark-grown higher plants the chlorophyll biosynthetic pathway stops at protochlorophyllide because the enzyme, NADPH-pchlide oxidoreductase (POR), responsible for the transformation of pchlide into chlorophyllide (chlide) requires light for catalysis. As a consequence, the transformation of pchlide into chlorophyllide can be triggered with illumination in dark-grown plants. The newly formed chlorophyllide-enzyme complexes undergo a spectral blue shift. The photoreduction and the blue spectral shift were studied in homogenates prepared from etiolated wheat leaves using fluorescence spectroscopy at pressures up to 400 MPa. The kinetic curves of photoreduction were fitted with the sum of two exponential functions. The activation volumes were calculated from the pressure dependence of the reaction rate constants of the exponential curves at 20 °C and 40 °C. The two activation volumes corresponding to the two processes linked to the photo-transformation of pchlide were 1.8±1.2 mL mol−1 and 2.9±0.5 mL mol−1 at 20 °C, and both activation volumes increased with increasing temperature. The blue shift showed strong pressure dependence below 100 MPa at 20 °C and was almost stopped at pressures higher than this value. The activation volume obtained for the pressure dependent phase of the blue shift was 43±11 mL mol−1. This activation volume decreased at 40 °C. The temperature dependence of the processes was analyzed at atmospheric pressure between 10 °C and 40 °C. The activation energy of the processes was calculated using Arrhenius plot. The activation energy of the blue shift was 98±20 kJ mol−1.
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