Journal
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
Volume 135, Issue 47, Pages 17869-17880Publisher
AMER CHEMICAL SOC
DOI: 10.1021/ja408230k
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Funding
- National Institutes of Health [DK046335]
- Skaggs Institute for Chemical Biology
- Lita Annenberg Hazen Foundation
- NRSA from the NHLBI [F32-HL099245]
- DOE Office of Biological and Environmental Research
- National Institute of General Medical Sciences [P41GM103393]
- National Center for Research Resources of the National Institutes of Health [P41RR001209]
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Small molecules that react selectively with a specific non-enzyme drug-target protein in a complex biological environment without displacement of a leaving group (tracelessly) are rare and highly desirable. Herein we describe the development of a family of fluorogenic stilbene-based vinyl amides and vinyl sulfonamides that covalently modify transthyretin (TTR) tracelessly. These small molecules bind selectively to TTR in complex biological environments and then undergo a rapid and chemoselective 1,4-Michael addition with the pK(a)-perturbed Lys-15 epsilon-amino group of TTR. Replacing the vinyl amide in 2 with the more reactive vinyl sulfonamide in 4 hastens the conjugation kinetics. X-ray cocrystallography verified the formation of the secondary amine bond mediating the conjugation in the case of 2 and 4 and confirmed the expected orientation of the stilbene within the TTR binding sites. Vinyl amide 2 and vinyl sulfonamide 4 potently inhibit TTR dissociation and amyloid fibril formation in vitro. The TTR binding selectivity, modification yield, and reaction chemoselectivity of vinyl sulfonamide 4 are good enough in human plasma to serve as a starting point for medicinal chemistry efforts. Moreover, vinyl sulfonamide 4 is fluorogenic: it exhibits minimal background fluorescence in complex biological environments, remains dark upon binding to TTR, and becomes fluorescent only upon reaction with TTR. The fluorogenicity of 4 was utilized to accurately quantify the native TTR concentration in Escherichia coli lysate using a fluorescence plate reader.
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