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. 2023 Jan;30(1):72-80.
doi: 10.1038/s41594-022-00862-z. Epub 2023 Jan 2.

Computational design of constitutively active cGAS

Quinton M Dowling  1   2 , Hannah E Volkman  3 , Elizabeth E Gray  3   4 , Sergey Ovchinnikov  2   5 , Stephanie Cambier  3 , Asim K Bera  2   6 , Banumathi Sankaran  7 , Max R Johnson  2   6 , Matthew J Bick  2   6   8 , Alex Kang  2   6 , Daniel B Stetson  3 , Neil P King  9   10
Affiliations

Computational design of constitutively active cGAS

Quinton M Dowling et al. Nat Struct Mol Biol. 2023 Jan.

Abstract

Cyclic GMP-AMP synthase (cGAS) is a pattern recognition receptor critical for the innate immune response to intracellular pathogens, DNA damage, tumorigenesis and senescence. Binding to double-stranded DNA (dsDNA) induces conformational changes in cGAS that activate the enzyme to produce 2'-3' cyclic GMP-AMP (cGAMP), a second messenger that initiates a potent interferon (IFN) response through its receptor, STING. Here, we combined two-state computational design with informatics-guided design to create constitutively active, dsDNA ligand-independent cGAS (CA-cGAS). We identified CA-cGAS mutants with IFN-stimulating activity approaching that of dsDNA-stimulated wild-type cGAS. DNA-independent adoption of the active conformation was directly confirmed by X-ray crystallography. In vivo expression of CA-cGAS in tumor cells resulted in STING-dependent tumor regression, demonstrating that the designed proteins have therapeutically relevant biological activity. Our work provides a general framework for stabilizing active conformations of enzymes and provides CA-cGAS variants that could be useful as genetically encoded adjuvants and tools for understanding inflammatory diseases.

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References

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