Biotinylated Indoles as Probes for Enzymes of Auxin Biosynthesis

In an attempt to provide tools for the detection of plant enzymes which use indole as a substrate [N. I. Rekoslavskaya, R. S. Bandurski (1994) Phytochemistry 35, 905-909; J. Normanly, J. P. Slovin, J. D. Cohen (1995) Plant Physiol. 107, 323-329], we prepared indole derivatives coupled to biotin via an oligopeptide spacer. With the indole moiety anchored to the active site of the enzyme, the biotin moiety would be detected by means of avidin coupled to a suitable indicator molecule. As an example, we here report on constructs 1 which contain a spacer of 3 or 4 concatenated beta-alanine moieties linked to the aliphatic nitrogen of 3-, 5-, or 6-(2-aminoethyl)indole. With the appropriate 2-aminoethyl indole as the starting compound, the syntheses were initiated by condensation with the hydroxysuccinimide ester of an N-protected oligo-(beta-alanine). The products were, after N-deprotection, reacted with the hydroxysuccinimide ester of biotin to yield the title compounds 1 which were obtained as colorless solids moderately soluble in water. Compounds 1 were characterized by HPLC-MS-FAB (glycerol matrix) which afforded the protonated ions [M + H]^+ at the expected amu values, thus confirming their elemental composition, as well as the corresponding daughter ions. Further confirmation of the structures was obtained from C-13 and H-1 NMR spectra which showed the signals of the 2-aminoethyl indole, biotin, and beta-alanine moieties at the expected chemical shift values. Constructs 1 were able to bind simultaneously to enzyme-conjugated avidin and to model indole-binding proteins, thus indicating their usefulness as a promising tool for future research.