Reversing open reading frames to create a pseudo-mirror image of life

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This project explores the consequences of reversing the order of amino acids in a protein. Reversing the order in which a protein is made has the potential to discover enhanced or completely new protein functions that can help improve daily life. Proteins that speed-up chemical reactions are routinely used in washing detergents, toothpaste, and fuel treatments, to name a few household items. These active proteins are taken from nature and can be improved to do their job better. However, with each improvement, further improvements become harder to attain. Reversing a protein recreates a natural protein in a new way that may be able to break through these barriers to further improvement. Research on these reverse proteins is being conducted with students that include members of underrepresented groups in science so that the next generation of scientists will be ready to better our lives with these active proteins. As the synthesis of new life forms opens up several lines of bioethical inquiry, these are addressed, first within the group, and then more broadly, with academic bioethics reports and with the general public through an established seminar series at a local museum. To make reverse proteins that function researchers at Baylor College of Medicine will use the machine learning algorithm AlphaFold2 to identify amino acid residues that prevent proper folding of reverse proteins. These identified amino acids are being changed in silico to determine if a different amino acid at that position improves folding. These computational predictions are also being tested experimentally to determine if they improve folding and function of the reverse protein. In a parallel approach, the functions of reverse proteins are being improved through directed evolution. These improved reverse proteins are being purified and tested for in vitro function, thermal stability, and suitability for protein crystallization so that their 3D structure can be determined. The findings from these experiments are improving understanding of how proteins fold and function and allowing scientists to create new proteins with enhanced or new functions.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.