Isoelectric focusing of pigment‐protein complexes solubilized from non‐irradiated and irradiated prolamellar bodies

Abstract

Preparative isoelectric focusing was employed to compare the association of protochlorophyllide and chlorophyllide with the enzyme NADPH‐protochlorophyllide oxidoreductase (PCR; EC 1.3.1.33). Photoactive protochlorophyllide‐PCR complexes were solubilized with 1‐O‐n‐octyl‐β‐d‐glucopyranoside from non‐irradiated prolamellar bodies of wheat (Triticum aestivum). Also, chlorophyllide‐PCR complexes were solubilized from prolamellar bodies irradiated under conditions either preventing or favouring a spectral shift of chlorophyllide to shorter wavelengths. Independently of the treatment prior to the solubilization, the pigments and the PCR focused together at pHs of 4 to 5. The results indicate that protochlorophyllide‐PCR complexes are conformationally similar to chlorophyllide‐PCR complexes. The results support the hypothesis that the spectral shift, referred to as the Shibata shift, reflects a breaking‐up of large chlorophyllide‐PCR aggregates to smaller chlorophyllide‐PCR units, rather than a dissociation of the chlorophyllide from the enzyme protein.