
are formed and some ketone remains unreduced. C: Treatment of the ketone with excess sodium borohydride (NaBH 4 ). Under these conditions, some ketone is reduced to afford diastereomers A1 and A2, some ketone undergoes dehydration, and a significant amount of the ketone remains unreduced. Figure S4 . LC--MS analysis of the conversion of seco A to seco B upon incubation with N--acetyl lysine (analysis of unreacted secosterols). Samples of N--acetyl lysine were incubated with excess seco A in pH 7.4 phosphate buffer:CH 3 CN (1:1) for varying times and subsequently reduced with NaBH 4 . (Some seco A formed adducts with N--acetyl lysine (data not shown); however five equivalents of seco A were used to allow for monitoring of the excess seco A over time. Buffer itself will catalyze the conversion, but it is greatly accelerated in the presence of N--acetyl lysine). Note that reduction with NaBH 4 results in a mixture of diastereomers (A1 and A3). SecoA readily dehydrates in the mass spectrometer so both m/z 423 and 405 were monitored. A: Authentic sample of (reduced) seco A. B: Incubation of seco A (5 equiv.) for 1 h at 37 °C in the presence of N--acetyl lysine (1 equiv.). C: Incubation of seco A (5 equiv.) for 3 h at 37 °C in the presence of N--acetyl lysine (1 equiv.). D: Incubation of seco A (5 equiv.) for 5 h at 37 °C in the presence of N--acetyl lysine (1 equiv.). Table S1 . Number of spectral counts and peptides identified in each SPE fraction analyzed by Myrimatch with 92% confidence. Spectra are filtered under stringent criteria: less than 2% Q value, a minimum of two distinct peptides and one additional peptide, and a minimum of two spectra per protein. All of the peptides identified in the 10 and 30 % fractions are unmodified HSA tryptic peptides. 75% of filtered spectra are eluted in 10-30% MeCN SPE fractions. Most of the modified peptides were contained in the 60% fraction listed in 3, 31.0, 29.8, 28.7, 28.0, 25.1, 24.0, 20.6, 18.7, 18.5, 15.9, 11.8, (minor) 69.3, 63.6, 56.2, 55.9, 51.2, 42.2, 37.2, 32.5, 29.7, 24.1, 21.9, 17.0, 11.7; HRMS (ESI) 6, 202.7, 84.7, 71.0, 68.1, 55.9, 54.2, 52.6, 46.8, 44.1, 42.5, 42.1, 39.8, 35.3, 34.8, 34.7, 34.0, 27.7, 27.6, 25.2, 25.0, 23.0, 18.7, 18.5, 17.5, 11.5 6, 84.7, 84.2, 68.1, 37.3, 63.9, 56.1, 55.5, 50.4, 45.5, 44.7, 44.2, 39.9, 39.6, 35.2, 35.0, 28.2, 27.9, 26.7, 25.1, 24.5, 21.5, 18.7, 18.6, 18.4, 12.5 4, 202.6, 147.7, 128.7, 85.0, 68.3, 56.2, 55.2, 48.0, 45.3, 42.6, 42.3, 40.0, 35.63, 35.56, 35.1, 34.0, 27.7, 25.4, 25.3, 24.9, 23.5, 19.0, 18.7, 18.0, 11.7 Synthesis of seco A triols. LiAlH 4 (0.7 mL, 1M/THF) was added to a solution of secoA (75 mg, 0.18 mmol) in THF.

After 30 min, the reaction mixture was quenched with 10% HCl and extracted with EtOAc. The organic layer was

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