# General and High-Throughput Small Molecule Screens and Selections for Metabolic Engineering

> **NIH NIH R01** · COLUMBIA UNIV NEW YORK MORNINGSIDE · 2021 · $666,491

## Abstract

Project Summary
The objective of this proposal is to create general, high-throughput assays that are modular and
broad in scope to overcome the current bottleneck in testing the enormous diversity required for
solving metabolic engineering problems. If successful, these technologies will enable powerful
directed evolution approaches to be routinely applied to the biosynthesis of natural products and
their analogs. Metabolic engineering involves library sizes of up to 1020, many orders of magnitude
beyond now routine protein engineering, because multiple genes not only in the biosynthetic
pathway but also in the host strain background must be optimized often synergistically. Yet, today
metabolic engineering is primarily performed by introducing just a few genetic modifications at a
time and then assaying the resulting strains by low throughput gas- and liquid-chromatography
mass spectrometry methods. Previous high-throughput assays employed in metabolic
engineering have been limited to unusual molecules, such as chromophores. Thus, here we apply
the concept of displacement of a competitor molecule from a protein receptor to develop two
general assays for metabolic engineering: the fluorescence polarization (FP) assay and the yeast
three-hybrid (Y3H) selection. The FP assay would be implemented as a first-generation, medium
throughput screen, as a step stone to the Y3H which would have higher throughput of greater
than 108. When carried out under the conditions of sexual reproduction with mutagenesis via
homologous recombination (HR), libraries of greater than 1020 can be searched. In collaboration
with the Tang laboratory (UCLA) and the Snyder laboratory (UChicago), we challenge our
technology with the metabolic engineering mission of increasing production titers of the fungal
anhydrotetracycline TAN-1612 and generating biologically active analogs in S. cerevisiae for
combating antibiotic resistance and applications beyond.

## Key facts

- **NIH application ID:** 10134372
- **Project number:** 5R01GM134293-02
- **Recipient organization:** COLUMBIA UNIV NEW YORK MORNINGSIDE
- **Principal Investigator:** VIRGINIA W CORNISH
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $666,491
- **Award type:** 5
- **Project period:** 2020-04-01 → 2024-02-29

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10134372

## Citation

> US National Institutes of Health, RePORTER application 10134372, General and High-Throughput Small Molecule Screens and Selections for Metabolic Engineering (5R01GM134293-02). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10134372. Licensed CC0.

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