Journal
NATURE COMMUNICATIONS
Volume 11, Issue 1, Pages -Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/s41467-020-17061-3
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Funding
- EU Marie Curie FP7 fellowships
- Biotechnology and Biological Sciences Research Council [BB/L002469/1, BB/M011194/1]
- Cambridge Service for Data Driven Discovery (CSD3)
- Engineering and Physical Sciences Research Council [EP/P020259/1]
- Science and Technology Facilities Council
- BBSRC [BB/L002469/1, 1644161] Funding Source: UKRI
- EPSRC [EP/P020259/1] Funding Source: UKRI
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Insertions and deletions (InDels) are frequently observed in natural protein evolution, yet their potential remains untapped in laboratory evolution. Here we introduce a transposon-based mutagenesis approach (TRIAD) to generate libraries of random variants with short in-frame InDels, and screen TRIAD libraries to evolve a promiscuous arylesterase activity in a phosphotriesterase. The evolution exhibits features that differ from previous point mutagenesis campaigns: while the average activity of TRIAD variants is more compromised, a larger proportion has successfully adapted for the activity. Different functional profiles emerge: (i) both strong and weak trade-off between activities are observed; (ii) trade-off is more severe (20- to 35-fold increased k(cat)/K-M in arylesterase with 60-400-fold decreases in phosphotriesterase activity) and (iii) improvements are present in k(cat) rather than just in K-M, suggesting adaptive solutions. These distinct features make TRIAD an alternative to widely used point mutagenesis, accessing functional innovations and traversing unexplored fitness landscape regions. Insertions/Deletions (InDels) remain an untapped source of protein diversity in laboratory evolution. The method TRIAD generates libraries of random variants with short in-frame InDels using transposons, allowing a comparison of their evolutionary potential with widely-used point mutant libraries.
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