NADPH:Protochlorophyllide Oxidoreductase (POR) : advanced insight into the catalytic mechanism of the two plant isozymes POR A and POR B

Garrone, Alessio GND

NADPH:protochlorophyllide oxidoreductase (POR), EC 1.3.1.33, is a key enzyme in the biosynthesis of chlorophyll in all organisms with oxygenic photosynthesis. POR catalyzes the reduction of protochlorophyllide (PChlide) to chlorophyllide (Chlide) by the absorption of light in the presence of the cofactor NADPH. In plants, two different isoenzymes are found, POR A and POR B. POR A is transiently active in etiolated plant seedlings, whereas POR is predominantly active throughout the greening process and in greened and adult plants. The two plant POR isoenzymes are compared with respect to their catalytic efficiency and their reaction mechanism in experiments using highly purified recombinant POR from barley (Hordeum vulgare L.). POR B shows a 6-fold higher catalytic efficiency (kcat/KM) but the same reaction mechanism when compared with POR A. The increased catalytic efficiency is attributed to a higher substrate binding affinity and a higher conformational flexibility of protein sites in the surrounding of the active centre. With the aim of gaining a deeper insight into the active-site architecture a 3-D model is predicted for POR B by using in silico homology modelling techniques. The prediction identifies His-138, Ser-164, Lys-216 and Tyr-299 as important residues for the correct positioning of the PChlide substrate within the active site. Moreover, the active site Tyr-212 and Lys-216 also play a key role in PChlide binding whereas Tyr-212 is also shown to take part in the proton transfer to the C(18) position of PChlide. The remaining residues within the catalytic motif, Lys-213, Asp214 and Ser-215 obviously create a polar interaction site, which is finely tuned by the cooperation of all three residues to facilitate the proper alignment of PChlide within the active site and to support a highly efficient photochemistry. Only small steric perturbations or changes in the hydrogen bonding tendency of these residues affect the photoreaction dramatically.

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Garrone, Alessio: NADPH:Protochlorophyllide Oxidoreductase (POR). advanced insight into the catalytic mechanism of the two plant isozymes POR A and POR B. Jena 2018.

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