Initial mutations direct alternative pathways of protein evolution

Area of Science:

• Evolutionary biology
• Molecular biology
• Biochemistry

Background:

• The predictability of evolutionary trajectories is debated, with epistasis potentially influencing both random and directed paths.
• Epistasis, non-additive interactions between mutations affecting fitness, can constrain mutation order or create contingency based on initial substitutions.
• Sign epistasis, where a mutation's fitness effect depends on its genetic background, strongly impacts adaptive pathways.

Purpose of the Study:

• To investigate how epistatic interactions between mutations shape alternative evolutionary pathways.
• To examine the role of epistasis in protein evolution using in vitro evolution of TEM-1 beta-lactamase.
• To understand the molecular basis of epistatic constraints on adaptive trajectories.

Main Methods:

• In vitro evolution of TEM-1 beta-lactamase to cefotaxime resistance.
• Analysis of diverse adaptive pathways in replicate experimental lines.
• Experimental evolution of alternative initial substitutions to assess their impact on subsequent adaptation.

Main Results:

• Most replicate lines evolved resistance through a common pathway involving three mutations in a fixed order, indicating epistatic constraints.
• A minority of lines followed divergent pathways, suggesting alternative initial substitutions led to different adaptive peaks.
• Negative sign epistasis, caused by decreased enzymatic activity and folding cooperativity, was identified between key mutations in common versus divergent pathways.

Conclusions:

• Epistasis significantly contributes to contingency in protein evolution.
• The selective consequences of random mutations are amplified by epistatic interactions.
• Understanding epistasis is crucial for predicting evolutionary trajectories and the development of antibiotic resistance.