Proteomic analysis of the etioplast inner membranes of wheat (Triticum aestivum) by two-dimensional electrophoresis and mass spectrometry

The proteome of the etioplast inner membranes (EPIM) of dark-grown wheat leaves (Triticum aestivum L.) was mapped as an essential part of studies on plastid differentiation. Proteins were separated by two-dimensional gel electrophoresis and analysed with mass spectrometry (MS). Over 200 protein spots were resolved and visualized by Coomassie blue staining. More than 100 spots were submitted for subsequent mass spectrometry analyses by matrix-assisted laser desorption ionization-time of flight (MALDI-ToF) MS, electrospray tandem MS (ESI-MS/MS) or liquid chromatography-mass spectrometry (LC-MS/MS). There were 46 identified spots, from which at least 21 different proteins were identified. Among these were FtsH proteases and the peptidyl-prolyl cis-trans isomerase TLP40, as well as chloroplast coupling factor subunits and extrinsic subunits of photosystem II (PSII). Of special interest is the NADPH:protochlorophyllide oxidoreductase (POR), which is the predominant protein of prolamellar bodies, where it accumulates in a highly stable ternary complex with protochlorophyllide and NADPH. This complex is known to play an important role in the formation and dispersal of prolamellar bodies. Five different isoforms of POR, with different pI values, were identified. We discuss the possibility of these isoforms being differently phosphorylated as part of the regulation of POR-pigment complexes. The proteome mapping of EPIM is a crucial step in the understanding of the light-dependent transition of etioplasts to chloroplasts, and provides a basis for functional studies on factors influencing the greening process.