Synthesis of enantiomerically pure cis- and trans-4-amino-1-oxyl-2,2,6,6-tetramethylpiperidine-3-carboxylic acid: a spin-labelled, cyclic, chiral beta-amino acid, and 3D-structural analysis of a doubly spin-labelled beta-hexapeptide

Amination of 3-carboxymethyl-l-oxyl-2,2,6,6-tetramethyl-4-piperidone (1) with either (R)- or (S)-alpha,-methylbenzylamine gave corresponding enamines 2. Whereas the reduction with. NaBH3CN/CH3COOH afforded predominantly a mixture of two possible cis diastereomers of 3, (1'R,3S,4S)1(1'R,3R,4R) or (VS,3R,4R)1(l'S,3S,4S), which could be separated by crystallisation of their HCl salts, the use of NaBH4/(CH3)(2)-HCOOH as the reducing agent resulted in a mixture of one trans- and one cis diastereomer of 3 (1'R,3S,4R)1(l'R,3R,4R) or (l'S,3R,4S)1(l'S,3S,4S) in varying proportions depending upon the conditions used. The stereochemistry of the four diastereomers of 3 was clearly established by X-ray diffraction analysis of one of them, combined with H-1 NMR spectroscopic studies after nitroxide reduction. Removal of the chiral auxiliary from the separated diastereomers of 3 by hydrogenation and regeneration of the nitroxide radical gave expected amino esters 4. A model beta-hexapeptide containing (3R,4S)beta-TOAC combined with (1S,2S)-2-aminocyclohexane carboxylic acid was synthesised by solution methods and its preferred conformation (3(14)-helix) was assessed by FTIR absorption, CD, and EPR spectroscopy.