Microwave-Mediated Reduction of Disulfide Bridges with Supported (Tris(2-carboxyethyl)phosphine) as Resin-Bound Reducing Agent

Abstract

We report on the synthesis and use of a new supported reagent consisting in tris(2-carboxyethyl)phosphine (TCEP) immobilized on hydrophilic PEG based resin beads. Used in conjunction with a 5 min microwave (MW) irradiation, "supported TCEP" reduced disulfide bridges in free thiols in peptides having two or more cysteine residues. Separation of reaction products from reducing agent was easily performed by simple filtration.

Reduction with tris(2-carboxyethyl)phosphine (TCEP) enables the use of an S-sulphonate protecting group for thiol-mediated bioconjugation

“…In this communication, we report the unprecedented use of TCEP (tris(2-carboxyethyl)phosphine) 12 and agarose-immobilized TCEP, an odourless and selective reducing reagent, 13 for the removal of the S-sulphonate protection releasing free thiols in neutral aqueous media. The compatibility of the deprotection conditions with a subsequent nucleophilic addition of the free thiols on a model maleimide substrate is demonstrated.…”

Protein Reduction and Dialysis‐Free Work‐Up through Phosphines Immobilized on a Magnetic Support: TCEP‐Functionalized Carbon‐Coated Cobalt Nanoparticles

“…Dialysis, the conventional TCEP removal procedure, is time‐consuming, has to be performed under non‐oxidizing conditions and causes loss of valuable proteins. Therefore, efforts for alternative solid support removal methods were considered: TCEP has previously been immobilized on polyethylene glycol (PEG) beads, but has only been shown effective for small peptides (e.g., mesethericin 3.8 kDa) and under microwave irradiation (at potentially problematic higher temperatures) . The commercially available TCEP bound on agarose Gel (Pierce™ Immobilized TCEP Disulfide Reducing Gel) has recently been introduced as a promising reducing agent also for larger proteins (at room temperature, using excess amounts of reducing agent).…”

Design of Nonhypertensive Conjugated Hemoglobins as Novel Resuscitation Fluids

“…The use of TCEP to deprotect the thiols at the distal end of the EAs with Hb is acceptable, as Hb does not contain any disulfide bonds. It should be noted that TCEP is an excellent reducing agent and can reduce the disulfide bonds of proteins (Miralles et al 2013;Jones et al 2012). Accordingly, it is recommended that when carrying out nonconservative EAF PEGylation protocol with disulfide-bonded proteins, for example albumin, milder deprotecting reagents like glutathione needs to be used to avoid the reduction of the intrinsic disulfide bonds of proteins.…”