File S1 - Transmembrane and Juxtamembrane Structure of αL Integrin in Bicelles

Includes Table S1, Figures S1–S3. Table S1. Structure statistics for the selected 20 structures of integrin αL TM domain. Figure S1. Expression and purification of integrin αL TM and β2 TM peptides. (A) SDS-PAGE of methanol-extracted (meOH) and HPLC-purified (HPLC) αL-TM and β2-TM peptides. (B) MALDI-TOF MS spectra of the HPLC fractions corresponding to αL TM and β2 TM peptides. The letters inside square brackets identify the charged species forming a particular peak: M, H, Na, K corresponds to peptide, proton, sodium ion, and potassium ions, respectively. The charge of each species is indicated outside the square bracket. Figure S2. Secondary structure of the peptide αL used, based on NOE connectivity. Sequential and medium-ranged NOE connectivity between residues are displayed as bands under the respective residues. The dαN(i,i+4) connectivities indicate the α-helical structure, extending from Leu-1065 to Val-1088. Figure S3. Secondary structure of integrin TM peptides. (A) FT-IR spectra of integrin TM α (upper) and β (lower) peptides reconstituted in DMPC lipid bilayers indicating that both TM peptides are almost completely α-helical. The amide I peak centered at 1657 cm−1 (blue) and the Fourier self-deconvolved spectra (red) are shown; (B) CD spectra of integrin TM peptides in DHPC-DMPC bicelles, showing minima at 209 and 222 nm, indicating a high proportion of α-helical form for both peptides; (C) Amide I and amide II regions of integrin TM peptides when bulk water was removed (red) and after D2O-saturated air exposure (blue). The decrease in the amide II area at ∼1550 cm−1 indicates H-D exchange. Figure S4. Effect of increasing concentration on the [1H-15N]-TROSY-HSQC spectrum of αL-TM. αL TM peptide at 0.1 mM (A) or 0.6 mM (B). αL was reconstituted into DHPC-DMPC bicelles (3% w/v, q = 0.3) buffered with 50 mM potassium phosphate at pH 6.5 and spectra were recorded at 305 K. (DOCX)