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

Helgeson, Luke A; Zelter, Alex; Riffle, Michael et al. (2018) Human Ska complex and Ndc80 complex interact to form a load-bearing assembly that strengthens kinetochore-microtubule attachments. Proc Natl Acad Sci U S A 115:2740-2745
Fong, Kimberly K; Zelter, Alex; Graczyk, Beth et al. (2018) Novel phosphorylation states of the yeast spindle pole body. Biol Open 7:
González, Delfina P; Lamb, Helen V; Partida, Diana et al. (2018) CBD-1 organizes two independent complexes required for eggshell vitelline layer formation and egg activation in C. elegans. Dev Biol 442:288-300
Basisty, Nathan B; Liu, Yuxin; Reynolds, Jason et al. (2018) Stable Isotope Labeling Reveals Novel Insights Into Ubiquitin-Mediated Protein Aggregation With Age, Calorie Restriction, and Rapamycin Treatment. J Gerontol A Biol Sci Med Sci 73:561-570
Brandsen, Benjamin M; Mattheisen, Jordan M; Noel, Teia et al. (2018) A Biosensor Strategy for E. coli Based on Ligand-Dependent Stabilization. ACS Synth Biol 7:1990-1999
Ma, Yuanhui; Yates 3rd, John R (2018) Proteomics and pulse azidohomoalanine labeling of newly synthesized proteins: what are the potential applications? Expert Rev Proteomics 15:545-554
Tseng, Boo Shan; Reichhardt, Courtney; Merrihew, Gennifer E et al. (2018) A Biofilm Matrix-Associated Protease Inhibitor Protects Pseudomonas aeruginosa from Proteolytic Attack. MBio 9:
Yates 3rd, John R (2018) Content Is King: Databases Preserve the Collective Information of Science. J Biomol Tech 29:1-3
DaRosa, Paul A; Harrison, Joseph S; Zelter, Alex et al. (2018) A Bifunctional Role for the UHRF1 UBL Domain in the Control of Hemi-methylated DNA-Dependent Histone Ubiquitylation. Mol Cell 72:753-765.e6
Xu, Yi; Ju, Ho-Jong; DeBlasio, Stacy et al. (2018) A Stem-Loop Structure in Potato Leafroll Virus Open Reading Frame 5 (ORF5) Is Essential for Readthrough Translation of the Coat Protein ORF Stop Codon 700 Bases Upstream. J Virol 92:

Showing the most recent 10 out of 372 publications