Mavor David, Barlow Kyle A, Asarnow Daniel, Birman Yuliya, Britain Derek, Chen Weilin, Green Evan M, Kenner Lillian R, Mensa Bruk, Morinishi Leanna S, Nelson Charlotte A, Poss Erin M, Suresh Pooja, Tian Ruilin, Arhar Taylor, Ary Beatrice E, Bauer David P, Bergman Ian D, Brunetti Rachel M, Chio Cynthia M, Dai Shizhong A, Dickinson Miles S, Elledge Susanna K, Helsell Cole V M, Hendel Nathan L, Kang Emily, Kern Nadja, Khoroshkin Matvei S, Kirkemo Lisa L, Lewis Greyson R, Lou Kevin, Marin Wesley M, Maxwell Alison M, McTigue Peter F, Myers-Turnbull Douglas, Nagy Tamas L, Natale Andrew M, Oltion Keely, Pourmal Sergei, Reder Gabriel K, Rettko Nicholas J, Rohweder Peter J, Schwarz Daniel M C, Tan Sophia K, Thomas Paul V, Tibble Ryan W, Town Jason P, Tsai Mary K, Ugur Fatima S, Wassarman Douglas R, Wolff Alexander M, Wu Taia S, Bogdanoff Derek, Li Jennifer, Thorn Kurt S, O'Conchúir Shane, Swaney Danielle L, Chow Eric D, Madhani Hiten D, Redding Sy, Bolon Daniel N, Kortemme Tanja, DeRisi Joseph L, Kampmann Martin, Fraser James S
Biophysics Graduate Group, University of California, San Francisco 94158, USA.
Bioinformatics Graduate Group, University of California, San Francisco 94158, USA.
Biol Open. 2018 Jul 23;7(7):bio036103. doi: 10.1242/bio.036103.
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, CaCl, 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.
尽管泛素（Ub）的一级蛋白质序列在进化过程中极其稳定，但在实验室进行的选择实验中，它对突变具有高度耐受性。我们提出，这种差异源于实验室培养条件下的适应性与进化时间尺度上不断变化的环境中的选择压力之间的差异。基于我们之前的工作（Mavor等人，2016年），我们使用深度突变扫描来确定十二种新化学物质（3-氨基-1,2,4-三唑、5-氟胞嘧啶、两性霉素B、氯化钙、浅蓝菌素、醋酸钴、甲萘醌、氯化镍、对氟苯丙氨酸、雷帕霉素、他莫昔芬和衣霉素）如何揭示泛素残基的新突变敏感性。总的来说，我们的实验确定了赖氨酸63的八个新敏化条件，并揭示了除丝氨酸57和谷氨酰胺62之外泛素中每个位置的一个敏化条件。通过确定不同化学约束下的泛素适应性景观，我们的工作有助于解决深度突变扫描实验与进化时间尺度上的序列保守性之间的不一致。
