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The importance of being knotted: effects of the C‐terminal knot structure on enzymatic and mechanical properties of bovine carbonic anhydrase II 1
Author(s) -
Alam Mohammad Taufiq,
Yamada Takafumi,
Carlsson Uno,
Ikai Atsushi
Publication year - 2002
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/s0014-5793(02)02693-5
Subject(s) - carbonic anhydrase , conformational isomerism , chemistry , circular dichroism , enzyme , carbonic anhydrase ii , knot (papermaking) , stereochemistry , crystallography , biochemistry , molecule , organic chemistry , materials science , composite material
In order to better understand the contribution of the knotted folding pattern to the enzymatic and mechanical properties of carbonic anhydrases, we replaced Gln‐253 of bovine carbonic anhydrase II with Cys, which allowed us to measure the mechanical strength of the protein against tensile deformation by avoiding knot tightening. The expressed protein, to our surprise, turned out to contain two conformational isomers, one capable of binding an enzymatic inhibitor and the other not, which led to their separation through affinity chromatography. In near‐ and far‐UV circular dichroism and fluorescence spectra, the separated conformers were very similar to each other and to the wild‐type enzyme, indicating that they both had native‐like conformations. We describe new evidence which supports the notion that the difference between the two conformers is likely to be related to the completeness of the C‐terminal knot formation.