A general strategy to construct small molecule biosensors in eukaryotes
Abstract
Biosensors for small molecules can be used in applications that range from metabolic engineering to orthogonal control of transcription. Here, we produce biosensors based on a ligand-binding domain (LBD) by using a method that, in principle, can be applied to any target molecule. The LBD is fused to either a fluorescent protein or a transcriptional activator and is destabilized by mutation such that the fusion accumulates only in cells containing the target ligand. We illustrate the power of this method by developing biosensors for digoxin and progesterone. Addition of ligand to yeast, mammalian, or plant cells expressing a biosensor activates transcription with a dynamic range of up to ~100-fold. We use the biosensors to improve the biotransformation of pregnenolone to progesterone in yeast and to regulate CRISPR activity in mammalian cells. This work provides a general methodology to develop biosensors for a broad range of molecules in eukaryotes.
Document Details
- Document Type
- Pub Defense Publication
- Publication Date
- Dec 29, 2015
- Source ID
- 10.7554/elife.10606
Entities
People
- Benjamin W Jester
- Christine E. Tinberg
- Daniel J. Mandell
- David Baker
- George M. Church
- June I. Medford
- Justin Feng
- Kevin J. Morey
- Mauricio S. Antunes
- Raj Chari
- Stanley Fields
- Xavier Rios
Organizations
- Colorado State University
- Defense Threat Reduction Agency
- Harvard Medical School
- Harvard University
- Howard Hughes Medical Institute
- National Institutes of Health
- National Science Foundation
- United States Department of Energy
- University of Washington