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Establishing catalytic activity on an artificial (βα) 8 ‐barrel protein designed from identical half‐barrels
Author(s) -
Sperl Josef M.,
Rohweder Bettina,
Rajendran Chitra,
Sterner Reinhard
Publication year - 2013
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/j.febslet.2013.06.022
Subject(s) - barrel (horology) , chemistry , atp synthase , protein engineering , isomerization , artificial enzyme , mutagenesis , biochemistry , enzyme , stereochemistry , catalysis , biophysics , biology , materials science , mutant , gene , composite material
It has been postulated that the ubiquitous (βα) 8 ‐barrel enzyme fold has evolved by duplication and fusion of an ancestral (βα) 4 ‐half‐barrel. We have previously reconstructed this process in the laboratory by fusing two copies of the C‐terminal half‐barrel HisF‐C of imidazole glycerol phosphate synthase (HisF). The resulting construct HisF‐CC was stepwise stabilized to Sym1 and Sym2, which are extremely robust but catalytically inert proteins. Here, we report on the generation of a circular permutant of Sym2 and the establishment of a sugar isomerization reaction on its scaffold. Our results demonstrate that duplication and mutagenesis of (βα) 4 ‐half‐barrels can readily lead to a stable and catalytically active (βα) 8 ‐barrel enzyme.