Tuning phenylalanine fluorination to assess aromatic contributions to protein function and stability in cells

Grace D Galles; Daniel T Infield; Colin J Clark; Marcus L Hemshorn; Shivani Manikandan; Frederico Fazan; Ali Rasouli; Emad Tajkhorshid; Jason D Galpin; Richard B Cooley; Ryan A Mehl; Christopher A Ahern

doi:10.1038/s41467-022-35761-w

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Tuning phenylalanine fluorination to assess aromatic contributions to protein function and stability in cells

Journal article

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Tuning phenylalanine fluorination to assess aromatic contributions to protein function and stability in cells

Grace D Galles, Daniel T Infield, Colin J Clark, Marcus L Hemshorn, Shivani Manikandan, Frederico Fazan, Ali Rasouli, Emad Tajkhorshid, Jason D Galpin, Richard B Cooley, …

Nature communications, Vol.14(1), 59

01/04/2023

DOI: 10.1038/s41467-022-35761-w

PMCID: PMC9813137

PMID: 36599844

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Abstract

The aromatic side-chains of phenylalanine, tyrosine, and tryptophan interact with their environments via both hydrophobic and electrostatic interactions. Determining the extent to which these contribute to protein function and stability is not possible with conventional mutagenesis. Serial fluorination of a given aromatic is a validated method in vitro and in silico to specifically alter electrostatic characteristics, but this approach is restricted to a select few experimental systems. Here, we report a group of pyrrolysine-based aminoacyl-tRNA synthetase/tRNA pairs (tRNA/RS pairs) that enable the site-specific encoding of a varied spectrum of fluorinated phenylalanine amino acids in E. coli and mammalian (HEK 293T) cells. By allowing the cross-kingdom expression of proteins bearing these unnatural amino acids at biochemical scale, these tools may potentially enable the study of biological mechanisms which utilize aromatic interactions in structural and cellular contexts.

Amino Acids - metabolism

Amino Acyl-tRNA Synthetases - metabolism

Escherichia coli - genetics

Escherichia coli - metabolism

Halogenation

HEK293 Cells

Humans

Phenylalanine - metabolism

RNA, Transfer - metabolism

Details

Title: Subtitle: Tuning phenylalanine fluorination to assess aromatic contributions to protein function and stability in cells
Creators: Grace D Galles - Oregon State University
Daniel T Infield - University of Iowa
Colin J Clark - University of Iowa
Marcus L Hemshorn - Oregon State University
Shivani Manikandan - University of Iowa
Frederico Fazan - University of Iowa
Ali Rasouli - University of Illinois Urbana-Champaign
Emad Tajkhorshid - University of Illinois Urbana-Champaign
Jason D Galpin - University of Iowa
Richard B Cooley - Oregon State University
Ryan A Mehl - Oregon State University
Christopher A Ahern - University of Iowa
Resource Type: Journal article
Publication Details: Nature communications, Vol.14(1), 59
DOI: 10.1038/s41467-022-35761-w
PMID: 36599844
PMCID: PMC9813137
NLM abbreviation: Nat Commun
ISSN: 2041-1723
eISSN: 2041-1723
Grant note: NS104617 / U.S. Department of Health & Human Services | NIH | National Institute of Neurological Disorders and Stroke (NINDS) GM144227 / U.S. Department of Health & Human Services | NIH | National Institute of General Medical Sciences (NIGMS) GM128420 / U.S. Department of Health & Human Services | NIH | National Institute of General Medical Sciences (NIGMS) GM104601 / U.S. Department of Health & Human Services | NIH | National Institute of General Medical Sciences (NIGMS) GM123455 / U.S. Department of Health & Human Services | NIH | National Institute of General Medical Sciences (NIGMS)
Language: English
Date published: 01/04/2023
Academic Unit: Molecular Physiology and Biophysics; Iowa Neuroscience Institute
Record Identifier: 9984357406002771

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