Abstract
We established and elaborated on a method to enrich the membrane proteome of mature pollen from economically relevant crop using the example of Solanum lycopersicum (tomato). To isolate the pollen protein fraction enriched in membrane proteins, a high salt concentration (750 mM of sodium chloride) was used. The membrane protein-enriched fraction was then subjected to shotgun proteomics for identification of proteins, followed by in silico analysis to annotate and classify the detected proteins.
Keywords: Membrane proteome, Pollen, Tomato, Proteomics
Background
As proper distribution of proteins and solutes between different cellular compartments or the insertion of newly-synthesized proteins into membranes is largely dependent on membrane proteins, the membrane proteome is central for maintenance of cellular and organellar homeostasis (Paul et al., 2013; 2014 and 2016a). Considering the importance of membrane proteins in general, these are also essential for pollen function and development (Paul et al., 2016b). Many global pollen proteomic studies have been performed in the past (Chaturvedi et al., 2013 and 2016); however, information about the intracellular distribution of proteins and the composition of the membrane proteome in pollen is rarely discussed (Pertl et al., 2009). One reason might be the low abundance and solubility of membrane proteins. Here we describe a protocol to isolate and analyze a protein fraction enriched in membrane proteins from mature pollen, which was established for tomato (Figure 1).Figure 1. Overview of the protocol for isolation of the membrane proteome of mature tomato pollen
Materials and Reagents
Equipment
Software
Procedure
Data analysis
Recipes
Acknowledgments
This protocol is adapted from (Paul et al., 2016a). This is an article of the SPOT-ITN consortium funded by Marie-Curie (EU Grant Agreement Number 289220) to E.S.
References
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