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Formation of local
asymmetric structures in the slow folding reaction
of human carbonic anhydrase II
Dick Andersson, IFM
Chemistry Linkopings University S-581 83 SWEDEN
In the study of protein folding we use human carbonic anhydrase
II
(HCA II) as a model protein. Circular dichroism (CD) and
fluorescence
have been used to monitor the folding process in engineered
tryptophan
mutants of HCA II. CD kinetic measurements on engineered
tryptophan
mutants of HCA II in the near- UV region have been used to probe
the
development of asymmetric environments around specific Trp
residues
during the refolding of HCA II. The formation of the active site
was
probed by the binding of a fluorescent sulfonamide inhibitor. So
far
three of the seven Trp residues in HCA II have been analyzed. The
three selected Trp residues are situated in different sites of
the
protein and they are the major contributors to the near-UV CD
spectrum. The aim of the study is to describe the formation of
specific native structures around individual sites during the
refolding process of HCA II. The refolding kinetics for the
studied
Trp residues shows that the asymmetric environment around these
Trp
residues is formed at different rates, showing that the formation
of
native-like tertiary structure occurs with varying rates in
different
regions of the protein.
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