ORBI  - EXPERIMENTAL PROCEDURE
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	Saccharomyces cerevisiae DBY8724 cells11 (MATa GAL2 ura3 bar1::URA3) were grown with aeration at 30 °C until O.D. 0.6-0.8 in either rich YPD medium (2 % yeast extract, 1 % peptone, 2 % glucose) or synthetic YMD minimal medium (0.7 % yeast nitrogen base without amino acids and ammonium sulfate (DIFCO Bacto), 2 % glucose, 5 g/L ammonium sulfate, and 20 mg/L uracil), pelleted, washed, resuspended in buffer (20 mM Tris-HCl, 100 mM NaCl) containing 1% protease inhibitor cocktail (Calbiochem, CA) and lysed with glass beads. Soluble protein extracts were diluted to 4 mg/ml into digestion buffer (50 mM Tris HCL pH 8.0, 1.0 M Urea, 2.0 mM CaCl2), denatured at 95 °C for 10 min, and digested with sequencing grade trypsin (Sigma, MO) at 37 °C for ~20 hours.  

	 For the mass spectrometry analysis, runs were performed on an LTQ-Orbitrap varying a range of parameters for optimization.  SCX salt steps were performed by injecting 10ul of Ammonium Chloride solutions of varying molarity, namely (0, 15, 60, 900) mM or (0, 20, 100, 900) mM in a 5% ACN, 0.1% Formic Acid background onto a strong cation exchange column (Thermo BioBasic-SCX 100mm X 0.180 mm ID) with a flow rate of 800 nl/min. Reverse phase chromatography was performed on a Thermo BioBasic-18 column 100mm X 0.10 mm ID running 38 nl/min for 90 or 120 minutes with varying ACN concentrations on a background of 0.1% Formic Acid.  The column eluent was nano-electro-sprayed at 1.95kV from a 10uM tip (New Objective). FTMS resolution was set at 60,000 or 100,000. Between 6 and 10 MS/MS spectra (using 1 or 3 microscans) were acquired per MS scan. Many other individual parameters were varied including exclusion list settings, charge state rejection, mono-isotopic precursor selection, minimum signal required for MS2 scanning, MS2 isolation width, and use of a mass lock (445.120025).  
     
     The probability of observing each protein and the total number of observed peptides were calculated using PeptideProphet and ProteinProphet, selecting proteins above a 5% false discovery rate (FDR) for protein identification threshold.