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21.11.7.3 Diselenide–Selenoester Ligation

DOI: 10.1055/sos-SD-121-00066

Malins, L. R.; Payne, R. J.Science of Synthesis Knowledge Updates, (20213226.

One of the key limitations of native chemical ligation (NCL) chemistry is the slow kinetics of the reaction at peptide thioesters bearing sterically hindered amino acids at the C-terminus; for example, ligation at valine and isoleucine can require a reaction time of over 48 hours and often does not proceed to completion due to competing thioester hydrolysis. Inspired by the NCL reaction, a solution to this problem was found by harnessing the superior electrophilicity of peptide selenoesters, specifically Se-aryl selenoesters, and the enhanced nucleophilicity of the selenol on the side chain of a selenocysteine (Sec) residue at the N-terminus of the other peptide fragment. Specifically, when a peptide 123 bearing a C-terminal phenyl selenoester is mixed with a peptide diselenide dimer 124 bearing an N-terminal selenocystine (the oxidized form of Sec) in aqueous denaturing ligation buffer, selenopeptide ligation products 125127 are generated in minutes, even at sterically hindered ligation junctions (Scheme 21).[‌128‌,‌129‌] This reaction, dubbed the additive-free diselenide–selenoester ligation (DSL), is performed in the absence of any reductive or thiol/selenol additives and, depending on the steric environment at the ligation junction and equivalents of peptide selenoester 123, leads to three ligation products in varying amounts: symmetrical diselenide 125, asymmetric diselenide 126, and product 127 bearing a selenoester linkage on the Sec used for ligation. All of these products are substrates for in situ deselenization (see Section 21.11.7.1.6) with tris(2-carboxyethyl)phosphine (TCEP) and dithiothreitol (DTT), which converts selenocysteine into alanine to afford single native polypeptide product 128. Ligation reactions can also be subjected to oxidative deselenization with TCEP and Oxone (see Section 21.11.7.1.6) to convert selenocysteine into serine at the ligation junction in products 129 (Scheme 21).[‌130‌] It should be noted that, mechanistically, the additive-free DSL reaction is thought to proceed by an initiation step that leads to the release of phenylselenolate from an equivalent of peptide selenoester 123.[‌131‌] This leads to diselenide exchange on a peptide diselenide dimer, resulting in a Sec-containing peptide that can enter an NCL-like reaction pathway (trans-selenoesterification followed by Se-to-N acyl shift).

Meeeee 88 Meeeeeee-Meee MMM Meeeeeee ee Me Meee Meeeeeeee Meeeeeeeeeeeee ee Meeeeeeee Meeeeeeeeeeeee[‌888‌‌888‌]

Meeeeeeee ee MMM eeeeeeeee ee e eeeeee ee eeeee eeeee eee eeee eeeeeeee eeeeeee eee eeeeeeeee ee eeeeeeeeeee eeeee eeee eee eeeeeeee eeeeeeeee eee eeeeee eeeeeeeeeeeee eeee eeeee-eeeee eeeeeee eeeeeeeee (MMMM) (Meeeee 88). Meeeeeeeeeee, MMM–eeeeeeeeeeeeee eeeeeeeee eee, ee eeee, eeee eeeeeeeeeeee eeeeeeee ee eeeeeeee-eeeeeeeeeeeeee eeeeeeee eeee 888,[‌888‌] eeeeeeee eeee 888,[‌888‌] eeeeeee 888,[‌888‌,‌888‌] eeeeeeeeeeeee 888,[‌888‌] eee eeeeeee 888.[‌888‌] MMM eeeeeeeee ee eeeeeeee eeeeeee eeeee eeeeeeee ee eee M-eeeeeeee eee ee eeeeeeeee ee eeeee eee eeee eeeeeee eeeeeeee eeeeeeeee eeeee; eeeeeee, eeeeeeee eeeeeeeee ee 8-eeeeeeeeeeeee 888 eeeeeee 88–88 eeeee ee eeeee eeeeeeeeee. Me eee eeee ee eeeeeee, eeeee eeeee eeeee (eeeeeeee eeee eeeeeeeeeeeeee) eeee eeee eeeeeeeeeeee eeeeeeee ee eee MMM-eeeee eeeeeeeee ee eeeeeee eeeeeee eeeeeeee eeeeeeee.[‌88‌,‌888‌] Me eeeeee ee eeeee eeee eeeee eeeeeeee eeeeee eee eeee ee eeee ee eeeeeeee MMM-eeee eeeeeeeee eeee eeeeeee eeeeeeeeee (eee Meeeeee 88.88.8.8.8).[‌888‌,‌888‌]

Meeeee 88 Meeeeeeee Meeeeeeeeee Meeee eee Meeeeeeeeeeee eeee Meee-MMMM eee Mee ee MMM–Meeeeeeeeeeeee Meeeeeeee[‌888‌‌888‌]

Mee ee eee eeeeeeeee eeeeeeeeee ee eeeeeee eeeeeeee eeeeeeeee ee eeee eeeeeeeee eee eeeeeee eeeeeeeee ee eeeeeeeeee eeeeeeeeeeeeee. Meee ee eee ee eee eeeeeeeeeee eeeeee ee eee eeee-eeeeeeee eeeeeeeeeeeeeeeeeee eeee eeee eeeee eee eee eeeeeeee eeeeeeeee. M eeeee eeeeeeeeeee ee eeee eeeeeeeeeeeee eeeeeeeeee ee eeee eeeeee eeeeeee-eeeeeee eeeeeeeee, eeee ee eeeeeeeee ee eeeeeeeeeee-eeeee eeeeeeee, eee eee eeeeeeeee eeeee eeee eeeeeeee eeeeeeeee, eee eeeeeee MMM, eeeeeee eee eee ee eeeeeeeeee[‌888‌,‌888‌,‌888‌] ee eeeeeeeeeeee eeee.[‌888‌‌888‌] Me eee eeeeeeee-eeee eeee, eee MMM eeeeeeee eeeee ee eeeeeeee ee eeeeeeeeeeeeee ee eeeeeeee eeeeeeee eeeee 888 μM. Meeeeee, ee eeeeeeeeeeeee MMMM ee e eeeeeeeeee eeeeeeeee eee eeeeeeee eeeeeeeeee ee e eeeeeee eeeeeeeee (eeeee eeeeeeee MMMM-eeeeeeee eeeeeeeeeeeeee; Meeeee 88), eeeeeeee eeeeeeeee eeeeeee eeeeeeeeeee, eeee eeee ee eeeeeeeee eeeeeeeeeeeeee ee eee eeeeeee eeeeeeeeee eeeee 888 eee eeeeeee eeeeeeeeeee 888 eeeeeeeee (Meeeee 88).[‌888‌] Meee eeeeeeee eee eeee eeeeee eee eeeeeeeee eeeeeeeeee–eeeeeeeeeee eeeeeeee (eMMM) eeeeeeee. Me eeeeeeeeee eee eeeeeeee eeeeeeee ee eeee ee eeeeee eeeeeeee 888, e eeeeeeeeeeeee eeeeeeeeeeeeee ee eeeeeeee eeee eeeeeee eee eeeee eeeeeeeeee ee eeeeeeeeeeeeee ee eeeeeee ee eee eeeeeeee eeeeeeee ee eeeeeeeeeeeeee ee eee eeeeeee eeeeeee ee eee ee 888 μM. Meee eee-eee eMMM–eeeeeeeeeeeeee eeeeeeeeeee eee eeee eeee eee eee eeeeeeeeee eeeeeeee ee e eeeeeeeee eeeeeee eee ee eeeeeeee eeeeeeee eeeeeeeeeee.[‌888‌]

Meeeee 88 Meeeeeeee Meeeeeeeee–Meeeeeeeeee Meeeeeee Meeeeeee ee Me Meee Meeeeeeeeeeeeeeeeee[‌888‌]

Meeeeeeeeeee Meeeeeeee

Meeeeee-Meeeeeeeee Meeeeeee 888; Meeeeee Meeeeeeee eee Meeeeeee-Meee MMM Meeeeeee ee Meeeeeeeeeee Meeeeeeeeeeeee:[‌888‌,‌888‌]

Meeeeeeee eeee eeeeeeeee eeeeeeeee ee 8–8 ee ee eeeeeee eeeeeeeeee eeeee 888. Meeeeee eeeeeeeeeee 888 (8.8–8.88 eeeee) eee eeeeeee eeeeeeeeee 888 (8.8 eeeee) eeee eeeeeeeee eeeeeeeeee ee eeeeeeee eeeeee (8 M eeeeeeeeeMMe, 888 eM eeeeee eeeeeeeee eeeeee; eM 8.8) ee e eeeeeeeeeeeee ee 88 eM eeee eeeeeee ee eeeeeee eeeeeeeeeee 888 eee 8 eM eeee eeeeeee ee eeeeeee eeeeeeeeee eeeee 888. Mee eeeee eeee eeeeeeee ee eeee ee eeeeeee eeeeeeeeeeeee ee 8.8 eM eeee eeeeeee ee eeeeeeeeeeeee eeeee 888 (8 eM eeee eeeeeee ee eee eeeeeee) eee 8 eM eeee eeeeeee ee eeeeeee eeeeeeeeeee 888, eee e eeeee eM ee eee eeeeeee ee 8.8–8.8. Mee eeeeeeee eeeeeeee eee eeeeeee ee eeeeeee eee 8–88 eee eeeeee eeeeeeee eeeeeeeeee, ee eeeeeeee ee MMMM/MM ee MMMM/MM. Me eee eeeee eeeeeee eeeeeeee eeee (ee. 8 eM eeee eeeeeee ee eeeeeeee eeeeeeee 888-888) eee eeeee ee eeeee eeeeee ee eeeeee. Mee eeeeee eee eeeeeeee eee eeeeee eee eeeeeee. Mee eeeeeee eeeee eee eeee eeeeeeee eeeee e eeeeeee ee eeeee. M eeee ee MMMM (8.88 M) eee MMM (88 eM) ee M8M eee eeeeeeee, eee eM eee eeeeeeee ee 8.8–8.8 eeee 8 M MeMM, eee eee eeee eee eeeeeeee. Mee eeeee eeeeeeee eeee eee eeeeeee eeee ee eeeee eeeeee ee eee eeee ee MMMM eee MMM (88 eeeee ee MMMM eee 8 eeeee ee MMM) ee eeee e eeeee eeeeeeeeeeeee ee eeeee 8.8 eM eeee eeeeeee ee eeeeeee eeeeeeee eeeeeeee 888-888 eee e eeeee eM ee 8.8–8. Mee eeee eee eeeeeeee ee ee eeeeeee eeeeee ee ee eee eeeeeeeee ee MM/MM eeeeeeee. Meeeeeeee eeeeeeeeee ee eee eeeeeeee (eeeeeeeee 8–88 e), eee eeee eee eeeeeeee eee eeeeeeeeeeeeeee eeeeeee-eeeee MMMM, eeeeeeeee e eeeeee eeeee ee 8.8% MMM ee M8M (eeeeeee M) eee 8.8% MMM ee MeMM (eeeeeee M). Meeeeee eeeeeee 888 eee eeeeeeee ee e eeeee eeeee eeeeeeeee eeeeeeeeeeeeee; eeeeeee eeeee (eeee 8 eeeee): 88%-eeeee.

Meeeeeee 888; Meeeeee Meeeeeeee eee Mee-Mee eMMM–Meeeeeeeeeeeeeeeeee:[‌888‌]

Meeeeeeee eeee eeeeeeeee eeeeeeeee ee 8–8 ee ee eeeeeee eeeeeeeeee eeeeee 888. M eeeeeeeeee ee 88 eM MMMM eee 88 eM eeeeeeee eeeeeeeeee ee eeeeeeee eeeeee (8 M eeeeeeeeeMMe, 8.8 M Mee-Meee, eM 8.8) eee eeeeeeee eee eeeeeeeee eee 88 eee. Mee eeeeeeeee eeee eee eeee ee eeeeeeeeee eeeeeeee eeeeeeee ee eeeeeeeeee eeeee eeeeeee 888 (88 μM eeee eeeeeee ee eee eeeee) eee eeeeeee eeeeee eeeeeeeeeee 888 (888 μM), eee eee eM eee eeeeeeee ee 8.8. Meeee eeeeeee ee eeeee ee 888 (8.8 eeeee eeee eeeeeee ee eee eeeee) eee 888 (8.8 eeeee) eeee eeeeeeee eee eeeeeeeee ee 88°M. Meee eeeeeeee eeeeeeee, ee eeeeeeeeee ee MMMM/MM eeeeeeee (eeeeeeeee 88–88 e), eee eeeeeee eee eeeeeeeee eeee Me8M (8 ×) eee eeeeeeee Me8M eee eeeeeee eeeee e eeeeee ee M8. Mee eeeeeeee eeee eee eeee eeeeeeee eeee ee eeeee eeeeee ee 888 eM MMMM eee 88 eM eeeeeeeeeee ee eeeeeeee eeeeee (8 M eeeeeeeeeMMe, 8.8 M Mee-Meee; eM 8.8) eee eeeeeeeeee ee e Meeeeee MMM-888 eeeeeeeeeeee (λ = 888 ee, 88 M) ee ee. Mee eeeeeeee eeee eee eeeeee eeee e eeee ee M88 eeeeee (88 ee) eee eeeeee eeeee eeee eeeeeee eee eeeeeee eeee 8.8% ee MMM (88 eM). Mee eeeeeeee eeeeeeee eee eeee eeeeee eeeee M8M/MeMM (8:8; 8 eM) eeeeeeeeee 8.8% MMM eeeeeeee ee MeMM (8 eM) eeeeeeeeee 8.8% MMM. Mee eeeeeeee eeeeeee eeee eeeeeeeeeee eee eee eeeeeeeee eeeeeee 888 eee eeeeeeee ee MMMM/MM. Mee eeeeeee ee eee eMMM–eeeeeeeeeeeeeeeeeee eeeeeeee eee eeee eeeeeeee ee eeeeeee-eeeee MMMM; eeeeeee eeeee (eeee 8 eeeee): 88–88%.

References


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