(27ae) Environmental Dependence of Collateral Fitness Effects

Mutational effects on proteins are commonly thought of in terms of how they affect the ability of the protein to perform its physiological function. Thus, when mutations deleteriously impact the growth rate of cells (i.e. fitness), effects on protein specific activity or protein abundance are usually expected. However, proteins must carry out their function in the complex milieu of the cell, where opportunities for something to go wrong abound. If a mutation causes a protein to decrease fitness through negatively affecting some cellular process unrelated to the physiological function of the protein, these effects are called collateral fitness effects. For example, a mutation might cause a protein to misinteract with an essential enzyme thereby slowing growth. We previously used deep mutational scanning to determine the collateral fitness effects of all single amino acid substitutions in TEM-1 β-lactamase by measuring fitness effects of mutations in the absence of β-lactam antibiotics. Over 42% of mutations caused deleterious collateral fitness effects. We selected over thirty such mutations for further study. All caused TEM-1 aggregation, suggesting that aggregation might be the cause of the deleterious effects. Here, we probe that hypothesis by asking how collateral fitness effects depend on the environment. We measured collateral fitness effects at 30°C in LB media, at 37°C in LB and M9 minimal media, and at 42°C in LB media. For select deleterious mutants we characterized how the different conditions affected the amount of soluble and aggregated TEM-1 and how that related to fitness effects. The results belie a simple relationship between aggregation and deleterious collateral fitness effects. For example, although both shifting from 37°C to 30°C in LB media and shifting from LB to minimal media at 37°C ameliorated the deleterious collateral fitness effects, aggregation typically did not decrease concomitantly. The results call in to question the hypothesis that deleterious collateral fitness effects are simply the result of the extent of protein aggregation.