Synthesis of N-Peptide-6-Amino-d-Luciferin Conjugates with Optimized Fragment Condensation Strategy

Abstract

The synthesis of peptide-luciferin conjugates has a pivotal role in the development of bioluminescent detection systems that are based on the determination of protease enzyme activity. This work describes the optimized synthesis of an N-peptide-6-amino-d-luciferin conjugate (Fmoc-Gly-Pro-6-amino-d-luciferin) with a simple fragment condensation method in adequate yields. Fmoc-Gly-Pro-6-amino-d-luciferin was produced from a previously synthesized Fmoc-Gly-Pro-OH and also previously synthesized 6-amino-2-cyanobenzothiazole with an optimized method, to which conjugate cysteine was added in an also improved way. The resulting conjugate was successfully used in a bioluminescent system, in vitro, demonstrating the applicability of the method. Graphical AbstractKeywords Bioluminescence · Protease activity · Aminoluciferin · Conjugate · Fragment condensation Electronic supplementary material The online version of this article (https ://doi.org/10.1007/s1098 9-018-9768-8) contains supplementary material, which is available to authorized users.

Solid-Phase Synthesis of Caged Luminescent Peptides via Side Chain Anchoring

“…17 Preliminary work by Kovacs et al revealed that loading of the carboxylic acid moiety of aLuc onto a solid support and subsequent elongation of the peptide chain was unsuccessful, due to the inherent instability of the thiazoline moiety of aLuc (Figure 1B). 18 The thiazoline ring is prone to oxidation resulting in the corresponding thiazole derivative, a well-known luciferase inhibitor. 19,20 Herein, we present an SPPS method for the synthesis of caged luminescent peptides starting with the side chain anchoring of either a lysine or an ornithine P 1 residue (Figure 1C).…”

“…The use of solid-phase peptide synthesis (SPPS) to construct the caged luciferins would drastically lower the number of steps in solution-phase, eliminate multiple purification steps, and especially allow for a parallel automated workflow . Preliminary work by Kovacs et al revealed that loading of the carboxylic acid moiety of aLuc onto a solid support and subsequent elongation of the peptide chain was unsuccessful, due to the inherent instability of the thiazoline moiety of aLuc (Figure B) . The thiazoline ring is prone to oxidation resulting in the corresponding thiazole derivative, a well-known luciferase inhibitor. , …”