Aryldithioethyloxycarbonyl (Ardec): A New Family of Amine Protecting Groups Removable under Mild Reducing Conditions and Their Applications to Peptide Synthesis

Abstract

The development of phenyldithioethyloxycarbonyl (Phdec) and 2-pyridyldithioethyloxycarbonyl (Pydec) protecting groups, which are thiol-labile urethanes, is described. These new disulfide-based protecting groups were introduced onto the epsilon-amino group of L-lysine; the resulting amino acid derivatives were easily converted into N alpha-Fmoc building blocks suitable for both solid- and solution-phase peptide synthesis. Model dipeptide(Ardec)s were prepared by using classical peptide couplings followed by standard deprotection protocols. They were used to optimize the conditions for complete thiolytic removal of the Ardec groups both in aqueous and organic media. Phdec and Pydec were found to be cleaved within 15 to 30 min under mild reducing conditions: i) by treatment with dithiothreitol or beta-mercaptoethanol in Tris.HCl buffer (pH 8.5-9.0) for deprotection in water and ii) by treatment with beta-mercaptoethanol and 1,8-diazobicyclo[5.4.0]undec-7-ene (DBU) in N-methylpyrrolidinone for deprotection in an organic medium. Successful solid-phase synthesis of hexapeptides Ac-Lys-Asp-Glu-Val-Asp-Lys(Ardec)-NH2 has clearly demonstrated the full orthogonality of these new amino protecting groups with Fmoc and Boc protections. The utility of the Ardec orthogonal deprotection strategy for site-specific chemical modification of peptides bearing several amino groups was illustrated firstly by the preparation of a fluorogenic substrate for caspase-3 protease containing the cyanine dyes Cy 3.0 and Cy 5.0 as FRET donor/acceptor pair, and by solid-phase synthesis of an hexapeptide bearing a single biotin reporter group.

A HTS Assay for the Detection of Organophosphorus Nerve Agent Scavengers

“…We and others [44] have already observed that such a thiol is stable up to pH 7.5, and that its spontaneous decomposition is pH-dependant, the decomposition being spontaneous above pH 8.0, which is consistent with our results described herein.…”

Discovery of a Dipeptide Epimerase Enzymatic Function Guided by Homology Modeling and Virtual Screening

“…Dipeptides for kinetic characterization were purchased when possible (e.g., Sigma, Bachem, Research Organics, Indofine, or MP Biochemicals), with the following exceptions: L-Ile-L-Phe was synthesized according to the procedure of Theodoropoulos and Craig (1955), and L-Lys-L-Phe was synthesized according to that of Lapeyre et al (2006). Syntheses for all other dipeptides are provided in Supplemental Data.…”

Biochemical Characterization of a Caspase-3 Far-red Fluorescent Probe for Non-invasive Optical Imaging of Neuronal Apoptosis

“…However, these probes have major limitations since they are unable to distinct apoptotic from necrotic cells, inasmuch as they penetrate into permeabilized necrotic cells and subsequently bind to phosphatidylserines that are 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 6 at the internal side of the cell membrane. More recently, other probes have been developed that directly target caspase-3 and become fluorescent only in the presence of active caspase-3, thus substantially increasing signal specificity (Bullok and Piwnica-Worms 2005;Lapeyre et al 2006). Functionality of these probes was demonstrated in human colon xenografts and in liver abscess mouse models (Bullok et al 2007), in isolated heart reperfusion injury (Pantos et al 2009) or after intra-dermal injection of caspase-3 (Zhang et al 2009).…”