Peptide Ligation at High Dilution via Reductive Diselenide-Selenoester Ligation

Abstract

Peptide ligation chemistry has revolutionized protein science by providing access to homogeneously modified peptides and proteins. However, lipidated polypeptides and integral membrane proteinsan important class of biomoleculesremain enormously challenging to access synthetically owing to poor aqueous solubility of one or more of the fragments under typical ligation conditions. Herein we describe the advent of a reductive diselenide-selenoester ligation (rDSL) method that enables efficient ligation of peptide fragments down to low nanomolar concentrations, without resorting to solubility tags or hybridizing templates. The power of rDSL is highlighted in the efficient synthesis of the FDA-approved therapeutic lipopeptide tesamorelin and palmitylated variants of the transmembrane lipoprotein phospholemman (FXYD1). Lipidation of FXYD1 was shown to critically modulate inhibitory activity against the Na+/K+ pump.

Traceless native chemical ligation of lipid-modified peptide surfactants by mixed micelle formation

“…Development of a lipid-facilitated NCL methodology that allows the rapid production of ligated polypeptides, using low concentrations of reactants 43 and in the absence of thiol additive catalysts, would be of considerable value. Herein, we describe a strategy for lipid-facilitated acceleration of NCL via micelle mixing ( Fig.…”

Challenges and Perspectives in Chemical Synthesis of Highly Hydrophobic Peptides

“…The deselenization reaction is further employable to other amino acids, such as aspartate and glutamate auxiliaries (Conibear et al, 2018) and is even employable to poorly soluble compounds. This is demonstrated by synthesis of the poorly soluble therapeutic lipopeptide tesamorelin and variants of the transmembrane lipoprotein phospholemman FXYD1 using this method and nanomolar concentrations circumventing the integration of solubilizing units (Chisholm et al, 2019). Broadening the ligation toolkit, the α-ketoacidhydroxylamine (KAHA) ligation route is also employable to poorly soluble and highly hydrophobic proteins such as IFITM3 (see Table 1) or the antibacterial cyclic AS-48 protein (Rohrbacher et al, 2017) making use of 5-oxaproline within acidic conditions.…”

Site‐Selective Modification of Peptides and Proteins via Interception of Free‐Radical‐Mediated Dechalcogenation

“…[37] The development of diselenide-selenoester ligation (DSL) employs both selenium-containing fragments to enable rapid, additive-free peptide ligation. [38][39][40][41][42][43] Protocols that facilitate the post-ligation conversion of internal Cys and Sec residues to alanine (Ala), via desulfurization and deselenization, respectively, [44,45] permit peptide ligation at sites that contain this more abundant residue. Further developments in this field include the utilization of non-proteinogenic thiolated and selenolated residues, which, when coupled with dechalcogenation, [46,47] grant access to a broad range of ligation junctions, dramatically enhancing the scope of the technique.…”

“…Further developments to this powerful method [33] include the use of Sec‐containing peptides [34–36] and selenoester peptides to accelerate the ligation reaction [37] . The development of diselenide‐selenoester ligation (DSL) employs both selenium‐containing fragments to enable rapid, additive‐free peptide ligation [38–43] . Protocols that facilitate the post‐ligation conversion of internal Cys and Sec residues to alanine (Ala), via desulfurization and deselenization, respectively, [44, 45] permit peptide ligation at sites that contain this more abundant residue.…”