Extending chemical perturbations of the ubiquitin fitness landscape in a classroom setting reveals new constraints on sequence tolerance
Mavor D, Barlow KA, Asarnow D, Birman Y, Britain D, Chen W, Green EM, Kenner LR, Mensa B, Morinishi LS, Nelson CA, Poss EM, Suresh P, Tian R, Arhar T, Ary BE, Bauer DP, Bergman ID, Brunetti RM, Chio CM, Dai SA, Dickinson MS, Elledge SK, Helsell CVM, Hendel NL, Kang E, Kern N, Khoroshkin MS, Kirkemo LL, Lewis GR, Lou K, Marin WM, Maxwell AM, McTigue PF, Myers-Turnbull D, Nagy TL, Natale AM, Oltion K, Pourmal S, Reder GK, Rettko NJ, Rohweder PJ, Schwarz DMC, Tan SK, Thomas PV, Tibble RW, Town JP, Tsai MK, Ugur FS, Wassarman DR, Wolff AM, Wu TS, Bogdanoff D, Li J, Thorn KS, O'Conchúir S, Swaney DL, Chow ED, Madhani HD, Redding S, Bolon DN, Kortemme T, DeRisi JL, Kampmann M, Fraser JSBiology Open, 2018Abstract:
Although the primary protein sequence of ubiquitin (Ub) is extremely stable over evolutionary time, it is highly tolerant to mutation during selection experiments performed in the laboratory. We have proposed that this discrepancy results from the difference between fitness under laboratory culture conditions and the selective pressures in changing environments over evolutionary timescales. Building on our previous work (Mavor et al., 2016), we used deep mutational scanning to determine how twelve new chemicals (3-Amino-1,2,4-triazole, 5-fluorocytosine, Amphotericin B, CaCl2, Cerulenin, Cobalt Acetate,
Menadione, Nickel Chloride, p-Fluorophenylalanine, Rapamycin, Tamoxifen, and Tunicamycin) reveal novel mutational sensitivities of ubiquitin residues. Collectively, our experiments have identified eight new sensitizing conditions for Lys63 and uncovered a sensitizing condition for every position in Ub except Ser57 and Gln62. By determining the ubiquitin fitness landscape under different chemical constraints, our work helps to resolve the inconsistencies between deep mutational scanning experiments and sequence conservation over evolutionary timescales.