Peptide fragment condensation and cyclisation using a subtilisin variant with improved synthesis over hydrolysis ratio

Peter Jan Leonard Mario Quaedflieg (Inventor), Timo Nuijens (Inventor), Jan Metske van der Laan (Inventor), Dirk Barend Janssen (Inventor), Ana Toplak (Inventor), Bian Wu (Inventor)

Research output: Patent

Abstract

The invention relates to a method for enzymatically synthesizing an (oligo)peptide, comprising coupling (a) an (oligo)peptide C-terminal ester or thioester and (b) an (oligo)peptide nucleophile having an N-terminally unprotected amine, wherein the coupling is carried out in a fluid comprising water, and wherein the coupling is catalyzed by a subtilisin BPN' variant or a homologue thereof, which comprises the following mutations compared to subtilisin BPN' represented by SEQUENCE ID NO: 2 or a homologue sequence thereof : - a deletion of the amino acids corresponding to positions 75-83; - a mutation at the amino acid position corresponding to S221, the mutation being S221C or S221selenocysteine; preferably a mutation at the amino acid position corresponding to P225 wherein the amino acid positions are defined according to the sequence of subtilisin BPN' represented by SEQUENCE ID NO: 2. Further, the invention relates to an enzyme suitable for use as a catalyst in a method of the invention.
Original languageEnglish
Patent numberWO2016056913
Priority date10/10/2014
Publication statusPublished - 14-Apr-2016

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