A Multicomponent Stapling Approach to Exocyclic Functionalized Helical Peptides: Adding Lipids, Sugars, PEGs, Labels, and Handles to the Lactam Bridge

Abstract

Peptide stapling is traditionally used to lock peptide conformations into α-helical structures using a variety of macrocyclization chemistries. In an endeavor to add a diversity-generating tool to this repertoire, we introduce a multicomponent stapling approach enabling the simultaneous stabilization of helical secondary structures and the exocyclic N-functionalization of the side chain-tethering lactam bridge. This is accomplished by means of a novel solid-phase methodology comprising, for the first time, the on-resin Ugi reaction-based macrocyclization of peptide side chains bearing amino and carboxylic acid groups. The exocyclic diversity elements arise from the isocyanide component used in the Ugi multicomponent stapling protocol, which allows for the incorporation of relevant fragments such as lipids, sugars, polyethylene glycol, fluorescent labels, and reactive handles. We prove the utility of such exocyclic reactive groups in the bioconjugation of a maleimide-armed lactam-bridged peptide to a carrier protein. The on-resin multicomponent stapling proved efficient for the installation of not only one, but also two consecutive lactam bridges having either identical or dissimilar N-functionalities. The easy access to helical peptides with a diverse set of exocyclic functionalities shows prospect for applications in peptide drug discovery and chemical biology.

Long-range PEG stapling: macrocyclization for increased protein conformational stability and resistance to proteolysis

“…22 These Cys-centric approaches are generally limited to side-chain/side-chain crosslinks; in contrast, other approaches facilitate stapling in both side-chain and backbone contexts. 23 For example, lactam staples can be prepared via conventional peptide coupling chemistry [24][25][26][27][28][29] or by diverse chemoselective strategies, including the Ugi reaction; 30,31 direct thioester aminolysis; 32 native chemical ligation; [33][34][35] KAHA ligation; 36 traceless Staudinger ligation; 37 and a variety of enzymatic methods. [38][39][40] Other creative stapling strategies include C-H activation; [41][42][43] the Petasis reaction; 44 the Glaser reaction; 45 oxime 46,47 or hydrazone 48 formation; the copper(I)catalyzed azide-alkyne cycloaddition (CuAAC); [49][50][51][52][53][54][55] and olefin metathesis.…”

Improved Stability and Tunable Functionalization of Parallel β‐Sheets via Multicomponent N‐Alkylation of the Turn Moiety

“…Previously, MCRs have been used to produce conformationally constrained macrocyclic peptides featuring stable protein secondary structures, such as α‐helices and reverse turns . However, this chemistry has never been employed for the construction of non‐cyclic β‐sheets.…”

Steroid diversification by multicomponent reactions

“…These protocols enabled the introduction not only of a steroidal moiety at the N -terminus but also of lipid and affinity tags (e.g., biotin) [64]. Importantly, the effective implementation of on-resin Ugi-4CRs paved the way for the subsequent development of multicomponent macrocyclizations permitting the introduction of PEGs, sugars and fluorescent labels at resin-linked peptides [66–67].…”