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 beta-strands in a dominating beta-structure that extends through the entire protein, is formed relatively late during refolding. In contrast, it was found that beta-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 beta-structure.