Improved Automatic Conversion for Use with a Liquid-Phase Sequenator

Abstract

The spinning cup sequenator designed by Edman and Begg in 1967 (1) incorporated automation of the coupling and cleavage reaction; conversion of the anilinothiazolinone (ATZ)-amino acid obtained at each cycle of Edman degradation to the corresponding phenylthiohydantoin (PTH)-amino acid was performed manually as a separate step using aqueous acid. The conversion step was automated for the spinning cup instrument by Wittman-Liebold (2,3) and subsequently adopted in other laboratories (4) using trifluoroacetic acid (TFA) as the conversion reagent. An automated conversion device was devised for a solid-phase sequencer by Birr (5). The advantages of automating the conversion step have been enumerated (2–8) and are summarized here: 1. The background levels of PTH-amino acids obtained during automated conversion are reproducible from cycle to cycle, although they gradually accumulate during the degradation at varying rates. 2. The yields for a specific PTH-amino acid are generally reproducible under a given set of conditions, in contrast to certain PTH-amino acids obtained following manual conversion where the yields vary depending on time elapsed prior to conversion. 3. As the automated system may be maintained secure from oxygen, it was predicted that improved yields would occur relative to manual conversion because of prevention of oxidative desulfuration of phenylthiocarbamyl (PTC)-amino acids.