# Biophysical mechanisms of proteomic and fitness effects of synonymous substitutions

> **NIH NIH R01** · HARVARD UNIVERSITY · 2020 · $322,977

## Abstract

Project Summary/Abstract
The overarching goal of the proposed research is to elucidate the mechanisms by which
synonymous mutations affect the abundance and turnover of synthesized proteins in cytoplasm
with ensuing effect on fitness. We determined that synonymous substitutions in the folA gene
encoding the essential E. coli enzyme Dihydrofolate Reductase (DHFR) have broad effects:
synonymous substitutions of rare codons to frequent ones at the 5' end of the nucleotide
sequence cause significant drop of the amount of mRNA produced while the ones near the 3'-
terminus result in depletion of soluble, active DHFR protein. We will broadly investigate the
relationship between DHFR folding thermodynamics and kinetics and its post-translation
abundance in cytoplasm for a broad coverage of sequence space including both missense and
synonymous mutations. We will focus on the role that protein quality control (specific proteases
and chaperones) play in amplifying or mitigating fitness effects of synonymous mutations and
determine the evolutionary paths by which E. coli recovers the fitness losses incurred by
specific synonymous mutations. Lessons from the DHFR study will be used as guidance for a
broader exploration on the level of complete genomes of several organisms from E. coli to
human with the aim to link conservation of specific synonymous codons to the position-specific
aspects of protein folding pathways. To this end we will apply our recent algorithm for structure-
based predictions of protein folding pathways to predict location of rare conserved codons
based on only the three-dimensional structure of the protein. Altogether these studies will
provide deep mechanistic insights into the origin of fitness effects of synonymous substitutions
and their evolutionary consequences. It will help to discern the forces of evolutionary selection
from sequence analysis data. Further, it will enable to identify the causal relationship between
mutations and disease phenotype – an unmet medical need in the era of data driven
approaches to development of new therapeutics.

## Key facts

- **NIH application ID:** 9962428
- **Project number:** 5R01GM124044-04
- **Recipient organization:** HARVARD UNIVERSITY
- **Principal Investigator:** EUGENE I SHAKHNOVICH
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $322,977
- **Award type:** 5
- **Project period:** 2017-09-19 → 2021-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9962428

## Citation

> US National Institutes of Health, RePORTER application 9962428, Biophysical mechanisms of proteomic and fitness effects of synonymous substitutions (5R01GM124044-04). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/9962428. Licensed CC0.

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