Abstract

In TR&D 1, we develop approaches to make complex changes to cells, and we generate biosensors to report on the effects of some of these changes. In the first aim, we establish strategies to rapidly and combinatorially up- or down-regulate a set of genes in a common pathway in order to improve the expression of a desired protein, and optimize these approaches using a vaccine protein that is commercially made in yeast. In the second aim, we devise new selections for use in chemostats and turbidostats to identify the effects of small changes in proteins, in promoters and in untranslated regions of mRNAs. Further, we modify the turbidostat platform to enable ramped selections, allowing feedback on temperature or drug concentration to optimize selections. In the third aim, we design new proteins that bind to a protein-protein interface and disrupt an interaction. These inhibitory proteins will allow us to assess the roles of individual members of large protein families that are difficult to analyze. Finally, in the fourth aim, we generate new biosensors to detect the levels of a small molecule and of a protein produced in yeast.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Biotechnology Resource Grants (P41)
Project #
5P41GM103533-23
Application #
9663945
Study Section
Special Emphasis Panel (ZRG1)
Project Start
Project End
Budget Start
2019-04-01
Budget End
2020-03-31
Support Year
23
Fiscal Year
2019
Total Cost
Indirect Cost

  Institution

Name
University of Washington
Department
Type
DUNS #
605799469
City
Seattle
State
WA
Country
United States
Zip Code
98195

  Publications

Jester, Benjamin W; Tinberg, Christine E; Rich, Matthew S et al. (2018) Engineered Biosensors from Dimeric Ligand-Binding Domains. ACS Synth Biol 7:2457-2467
Kim, Yeoun Jin; Sweet, Steve M; Egertson, Jarrett D et al. (2018) Data-independent acquisition mass spectrometry to quantify protein levels in FFPE tumor biopsies for molecular diagnostics. J Proteome Res :
Keich, Uri; Noble, William Stafford (2018) Controlling the FDR in imperfect matches to an incomplete database. J Am Stat Assoc 113:973-982
Louka, Panagiota; Vasudevan, Krishna Kumar; Guha, Mayukh et al. (2018) Proteins that control the geometry of microtubules at the ends of cilia. J Cell Biol 217:4298-4313
Searle, Brian C; Pino, Lindsay K; Egertson, Jarrett D et al. (2018) Chromatogram libraries improve peptide detection and quantification by data independent acquisition mass spectrometry. Nat Commun 9:5128
Payea, Matthew J; Sloma, Michael F; Kon, Yoshiko et al. (2018) Widespread temperature sensitivity and tRNA decay due to mutations in a yeast tRNA. RNA 24:410-422
Seixas, Adriana; Alzugaray, María Fernanda; Tirloni, Lucas et al. (2018) Expression profile of Rhipicephalus microplus vitellogenin receptor during oogenesis. Ticks Tick Borne Dis 9:72-81
Luan, Qing; Zelter, Alex; MacCoss, Michael J et al. (2018) Identification of Wiskott-Aldrich syndrome protein (WASP) binding sites on the branched actin filament nucleator Arp2/3 complex. Proc Natl Acad Sci U S A 115:E1409-E1418
Smukowski Heil, Caiti; Burton, Joshua N; Liachko, Ivan et al. (2018) Identification of a novel interspecific hybrid yeast from a metagenomic spontaneously inoculated beer sample using Hi-C. Yeast 35:71-84
Ziegler, Yvonne S; Moresco, James J; Tu, Patricia G et al. (2018) Proteomic analysis identifies highly expressed plasma membrane proteins for detection and therapeutic targeting of specific breast cancer subtypes. Clin Proteomics 15:30

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