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Peptiligase, an Enzyme for Efficient Chemoenzymatic Peptide Synthesis and Cyclization in Water
Author(s) -
Toplak Ana,
Nuijens Timo,
Quaedflieg Peter J. L. M.,
Wu Bian,
Janssen Dick B.
Publication year - 2016
Publication title -
advanced synthesis and catalysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.541
H-Index - 155
eISSN - 1615-4169
pISSN - 1615-4150
DOI - 10.1002/adsc.201600017
Subject(s) - chemistry , peptide , combinatorial chemistry , bacillus subtilis , peptide synthesis , hydrolysis , enzyme , chemical ligation , stereochemistry , substrate (aquarium) , artificial enzyme , native chemical ligation , organic chemistry , biochemistry , cysteine , genetics , oceanography , bacteria , biology , geology
Abstract We describe a novel, organic cosolvent‐stable and cation‐independent engineered enzyme for peptide coupling reactions. The enzyme is a variant of a stable calcium‐independent mutant of subtilisin BPN′, with the catalytic Ser212 mutated to Cys and Pro216 converted to Ala. The enzyme, called peptiligase, catalyzes exceptionally efficient peptide coupling in water with a surprisingly high synthesis over hydrolysis (S/H) ratio. The S/H ratio of the peptide ligation reaction is correlated to the length of the peptide substrate and proved to be >100 for the synthesis of a 13‐mer peptide, which corresponds to >99% conversion to the ligated peptide product and <1% hydrolytic side‐reaction. Furthermore, peptiligase does not require a particular recognition motif resulting in a broadly applicable and traceless peptide ligation technology. Peptiligase is very robust, easy to produce in Bacillus subtilis , and its purification is straightforward. It shows good activity and stability in the presence of organic cosolvents and chelating or denaturing agents, enabling the ligation of poorly soluble (hydrophobic) or folded peptides. This enzyme could be useful for the (industrial) synthesis of diverse (pharmaceutical) peptides. In addition, peptiligase is able to efficiently catalyze head‐to‐tail peptide cyclization reactions.