Synthesis of Bifacial Peptide Nucleic Acids with Diketopiperazine Backbones

Abstract

We report a synthesis of bifacial peptide nucleic acids (bPNAs) with novel diketopiperazine (DKP) backbones that display unnatural melamine (M) bases, as well as native bases. To examine the structure–function scope of DKP bPNAs, we synthesized a set of bPNAs by using diaminopropionic acid, diaminobutyric acid, ornithine, and lysine derivatives to display the base-tripling motifs, which result in one, two, three, or four carbons linking the alpha carbon to the side-chain amine. Thermal denaturation of DNA hybrids with these bPNAs revealed that the optimal side-chain linkage was four carbons, corresponding to the lysine derivative. Accordingly, monomers displaying two bases per side-chain were prepared through double reductive alkylation of the ε-amine of Fmoc-lysine with acetaldehyde derivatives of adenine, cytidine, uridine, and melamine. With these building blocks in hand, DKP bPNAs were prepared to display a combination of native and synthetic (melamine) bases. Preliminary melting studies indicate binding signatures of cytidine- and melamine-displaying bPNAs to T-rich DNAs of noncanonical structure, though full characterization of this behavior is ongoing. The convenient and potentially scalable method described enables rapid access to DNA-binding scaffolds of low (<1 kD) molecular weight and previously established cell permeability. We expect that this straightforward and efficient approach to nucleic acid binders will enable studies on noncanonical nucleic acid hybridization.

Key words

peptide nucleic acids - diketopiperazines - DNA binding - triplexes

Publication History

Received: 27 February 2022

Accepted after revision: 18 March 2022

Accepted Manuscript online:
18 March 2022

Article published online:
28 April 2022

• References and Notes

• 1 Whitesides GM, Simanek EE, Mathias JP, Seto CT, Chin D, Mammen M, Gordon DM. Acc. Chem. Res. 1995; 28: 37
• 2 Zeng Y, Pratumyot Y, Piao X, Bong D. J. Am. Chem. Soc. 2012; 134: 832
• 3 Piao X, Xia X, Mao J, Bong D. J. Am. Chem. Soc. 2015; 137: 3751
• 4 Xia X, Piao X, Bong D. J. Am. Chem. Soc. 2014; 136: 7265
• 5 Xia X, Piao X, Fredrick K, Bong D. ChemBioChem 2013; 15: 31
• 6 Piao X, Xia X, Bong D. Biochemistry 2013; 52: 6313
• 7 Miao S, Liang Y, Marathe I, Mao J, DeSantis C, Bong D. J. Am. Chem. Soc. 2019; 141: 9365
• 8 Liang Y, Mao J, Bong D. Methods Enzymol. 2019; 623: 151
• 9 Liang Y, Miao S, Mao J, DeSantis C, Bong D. Biochemistry 2020; 59: 2410
• 10 Mao J, DeSantis C, Bong D. J. Am. Chem. Soc. 2017; 139: 9815
• 11 Mao J, Bong D. Synlett 2015; 26: 1581
• 12 Zhou Z, Xia X, Bong D. J. Am. Chem. Soc. 2015; 137: 8920
• 13 Miao S, Liang Y, Rundell S, Bhunia D, Devari S, Munyaradzi O, Bong D. Biopolymers 2021; 112: e23399
• 14 Avakyan N, Greschner AA, Aldaye F, Serpell CJ, Toader V, Petitjean A, Sleiman HF. Nat. Chem. 2016; 8: 368
• 15 Li Q, Zhao J, Liu L, Jonchhe S, Rizzuto FJ, Mandal S, He H, Wei S, Sleiman HF, Mao H, Mao C. Nat. Mater. 2020; 19: 1012
• 16 Cafferty BJ, Fialho DM, Khanam J, Krishnamurthy R, Hud NV. Nat. Commun. 2016; 7: 11328
• 17 Lange RF. M, Beijer FH, Sijbesma RP, Hooft RW. W, Kooijman H, Spek AL, Kroon J, Meijer EW. Angew. Chem. Int. Ed. 1997; 36: 969
• 18 Arambula JF, Ramisetty SR, Baranger AM, Zimmerman SC. Proc. Natl. Acad. Sci. U.S.A. 2009; 106: 16068
• 19 Liang Y, Miao S, Mao J, Devari S, Gonzalez M, Bong D. Chem. Eur. J. 2022; 28: e202103616
• 20 Zhou Z, Bong D. Langmuir 2013; 29: 144
• 21 Ma M, Bong D. Langmuir 2010; 27: 1480
• 22 Ma M, Paredes A, Bong D. J. Am. Chem. Soc. 2008; 130: 14456
• 23 Ma M, Gong Y, Bong D. J. Am. Chem. Soc. 2009; 131: 16919
• 24 Miao S, Bhunia D, Devari S, Liang Y, Munyaradzi O, Rundell S, Bong D. ACS Chem. Biol. 2021; 16: 1600
• 25 Xia X, Zhou Z, DeSantis C, Rossi JJ, Bong D. ACS Chem. Biol. 2019; 14: 1310
• 26 Munyaradzi O, Rundell S, Bong D. ChemBioChem 2022; in press DOI: 10.1002/cbic.202100707.
• 27 Rundell S, Munyaradzi O, Bong D. Biochemistry 2022; 61: 85
• 28 Bong DT, Clark TD, Granja JR, Ghadiri MR. Angew. Chem. Int. Ed. 2001; 40: 988
• 29 Veatch WR, Fossel ET, Blout ER. Biochemistry 1974; 13: 5249
• 30 Borthwick AD. Chem. Rev. 2012; 112: 3641
• 31 Heemstra JM, Liu DR. J. Am. Chem. Soc. 2009; 131: 11347
• 32 Sun X, Rai R, MacKerell AD. Jr, Faden AI, Xue F. Tetrahedron Lett. 2014; 55: 1905
• 33 DKPs 1a–j; General Procedure The appropriate Fmoc-amino acid 3 (1.0 equiv.) was dissolved in NMP at RT. HBTU (1.0 equiv) and Et₃N (5.0 equiv) were added and the mixture was stirred for 10 min before the addition of the appropriate amino acid ester 2 (1.0 equiv). The resulting mixture was stirred for 24 h at 60–80 °C while the reaction was monitored by HPLC and mass spectroscopy. The mixture then was cooled to RT and the solvent was removed under reduced pressure. A mixture of 95% water, 5% MeCN, and 0.07% TFA (10 mL; HPLC solvent A) was added and the mixture was sonicated for 15 mins then filtered through a 0.5-micron filter. The DKP product was then purified by preparative HPLC.
• 34 Hyrup B, Nielsen PE. Bioorg. Med. Chem. 1996; 4: 5
• 35 Appella DH. Curr. Opin. Chem. Biol. 2009; 13: 687
• 36 Pokorski JK, Witschi MA, Purnell BL, Appella DH. J. Am. Chem. Soc. 2004; 126: 15067
• 37 Englund EA, Appella DH. Angew. Chem. Int. Ed. 2007; 46: 1414
• 38 Vilaivan T. Acc. Chem. Res. 2015; 48: 1645