Separation and identification of the light harvesting proteins contained in grana and stroma thylakoid membrane fractions

Journal of Chromatography. a
A M TimperioL Zolla

Abstract

This paper presents the results of a study performed to develop a rapid and straightforward method to resolve and simultaneously identify the light-harvesting proteins of photosystem I (LHCI) and photosystem II (LHCII) present in the grana and stroma of the thylakoid membranes of higher plants. These hydrophobic proteins are embedded in the phospholipid membrane, and their extraction usually requires detergent and time consuming manipulations that may introduce artifacts. The method presented here makes use of digitonin, a detergent which causes rapid (within less than 3 min) cleavage of the thylakoid membrane into two subfractions: appressed (grana) and non-appressed (stroma) membranes, the former enriched in photosystem II and the latter containing mainly photosystem I. From these two fractions identification of the protein components was performed by separating them by reversed-phase high-performance liquid chromatography (RP-HPLC) and determining the intact molecular mass by electrospray ionization mass spectrometry (ESI-MS). By this strategy the ion suppression during ESI-MS that normally occurs in the presence of membrane phospholipids was avoided, since RP-HPLC removed most phospholipids from the analytes. Consequently, ...Continue Reading

References

Aug 4, 1970·Biochimica Et Biophysica Acta·P V SaneR B Park
Feb 8, 1994·Biochimica Et Biophysica Acta·S Jansson
Jul 4, 2001·Trends in Plant Science·A HaldrupH V Scheller
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Nov 18, 2003·The Journal of Biological Chemistry·Suping Zhang, Henrik Vibe Scheller

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Citations

May 6, 2008·Journal of the American Society for Mass Spectrometry·Dmitry GaletskiyMichael Przybylski
Jan 29, 2009·Journal of Separation Science·Zhaowei ZhangBi-Feng Liu
Dec 15, 2004·Phytochemical Analysis : PCA
Sep 3, 2004·The Journal of Biological Chemistry·Frank KrauseNorbert A Dencher

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