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Folding around the C‐terminus of human carbonic anhydrase II Kinetic characterization by use of a chemically reactive SH‐group introduced by protein engineering
Author(s) -
Freskgård Per-Ola,
Carlsson Uno,
Mårtensson Lars-Göran,
Jonsson Bengt-Harald
Publication year - 1991
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 - 10.1016/0014-5793(91)80922-p
Subject(s) - carbonic anhydrase , chemistry , protein folding , folding (dsp implementation) , protein engineering , biochemistry , carbonic anhydrase ii , characterization (materials science) , kinetic energy , enzyme , biophysics , biology , materials science , nanotechnology , engineering , electrical engineering , physics , quantum mechanics
We are characterizing the process of refolding of the enzyme human carbonic anhydrase II from the denatured state in guanidine hydrochloride. To describe the folding in defined parts of the protein we use protein engineering to introduce cysteine residues as unique chemically reactive probes. The accessibility of the cysteine SH‐group to the alkylating reagent iodoacetate, at different stages during refolding, is used to give a kinetic description of the folding process. The structuration of the C‐terminal part of the polypeptide chain, which is involved in a unique ‘knot’ topology, was investigated. Our results show that the structure around the C‐terminal, composed of the outermost β‐strands in a dominating β‐structure that extends through the entire protein, is formed relatively late during refolding. In contrast, it was found that β‐strands located in the interior of the protein were structured very rapidly. The final native structure is formed in a process that is slower than those observed for formation of β‐structure.