Extending chemical perturbations of the ubiquitin fitness landscape in a classroom setting reveals new constraints on sequence tolerance

David Mavor(University of California, San Francisco), Kyle A. Barlow(University of California, San Francisco), Daniel Asarnow(University of California, San Francisco), Yuliya Birman(University of California, San Francisco), Derek Britain(University of California, San Francisco), Weilin Chen(University of California, San Francisco), Evan M. Green(University of California, San Francisco), Lillian R. Kenner(University of California, San Francisco), Bruk Mensa(University of California, San Francisco), Leanna S. Morinishi(University of California, San Francisco), Charlotte Nelson(University of California, San Francisco), Erin M. Poss(University of California, San Francisco), Pooja Suresh(University of California, San Francisco), Ruilin Tian(University of California, San Francisco), Taylor Arhar(University of California, San Francisco), Beatrice Ary(University of California, San Francisco), David Bauer(University of California, San Francisco), Ian D. Bergman(University of California, San Francisco), Rachel M. Brunetti(University of California, San Francisco), Cynthia M. Chio(University of California, San Francisco), Shizhong Dai(University of California, San Francisco), Miles Sasha Dickinson(University of California, San Francisco), Susanna K. Elledge(University of California, San Francisco), Cole Helsell(University of California, San Francisco), Nathan L. Hendel(University of California, San Francisco), Emily L. Kang(University of California, San Francisco), Nadja Kern(University of California, San Francisco), Matvei Khoroshkin(University of California, San Francisco), Lisa L. Kirkemo(University of California, San Francisco), Greyson R. Lewis(University of California, San Francisco), Kevin Lou(University of California, San Francisco), Wesley M. Marin(University of California, San Francisco), Alison M. Maxwell(University of California, San Francisco), Peter F. McTigue(University of California, San Francisco), Douglas Myers-Turnbull(University of California, San Francisco), Tamas L Nagy(University of California, San Francisco), Andrew M. Natale(University of California, San Francisco), Keely Oltion(University of California, San Francisco), Sergei Pourmal(University of California, San Francisco), Gabriel K. Reder(University of California, San Francisco), Nicholas J. Rettko(University of California, San Francisco), Peter J. Rohweder(University of California, San Francisco), Daniel M. C. Schwarz(University of California, San Francisco), Sophia K. Tan(University of California, San Francisco), Paul V. Thomas(University of California, San Francisco), Ryan W. Tibble(University of California, San Francisco), Jason P. Town(University of California, San Francisco), Mary K. Tsai(University of California, San Francisco), Fatima S. Ugur(University of California, San Francisco), Douglas R. Wassarman(University of California, San Francisco), Alexander M. Wolff(University of California, San Francisco), Taiasean Wu(University of California, San Francisco), Derek Bogdanoff(University of California, San Francisco), Jennifer Li(University of California, Davis), Kurt S. Thorn(University of California, San Francisco), Shane Ó’Conchúir(QB3), Danielle L. Swaney(QB3), Eric D. Chow(University of California, San Francisco), Hiten D. Madhani(University of California, San Francisco), Sy Redding(University of California, San Francisco), Daniel N. Bolon(University of Massachusetts Chan Medical School), Tanja Kortemme(QB3), Joseph L. DeRisi(University of California, San Francisco), Martin Kampmann(University of California, San Francisco), James S. Fraser(QB3)
Biology Open
July 15, 2018
10.1242/bio.036103
Cited by 29Open Access
Full Text

Abstract

ABSTRACT 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.

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