Characterization of folding intermediates during urea-induced denaturation of human carbonic anhydrase II

Wahiduzzaman, Mohammad Aasif Dar, Md Anzarul Haque, Danish Idrees, Md Imtaiyaz Hassan, Asimul Islam, Faizan Ahmad

Research output: Contribution to journalArticlepeer-review

25 Scopus citations


Knowledge of folding/unfolding pathway is fundamental basis to study protein structure and stability. Human carbonic anhydrase II (HCAII) is a ∼29 kDa, β-sheet dominated monomeric protein of 259 amino acid residues. In the present study, the urea-induced denaturation of HCAII was carried out which was a tri-phasic process, i.e., N (native) ↔ XI ↔ XII ↔ D (denatured) with stable intermediates XI and XII populated around 2 and 4 M urea, respectively. The far-UV CD was used to characterize the intermediate states (XI and XII) for secondary structural content, near-UV CD for tertiary structure, dynamic light scattering for hydrodynamic radius and ANS fluorescence spectroscopy for the presence of exposed hydrophobic patches. Based on these experiments, we concluded that urea-induced XI state has characteristics of molten globule state while XII state bears characteristics features of pre-molten globule state. Characterization of the intermediates on the folding pathway will contribute to a deeper understanding of the structure-function relationship of HCAII. Furthermore, this system may provide an excellent model to study urea stress and the strategies adopted by the organisms to combat such a stress.

Original languageEnglish (US)
Pages (from-to)881-887
Number of pages7
JournalInternational Journal of Biological Macromolecules
StatePublished - Feb 1 2017
Externally publishedYes


  • Human carbonic anhydrase II
  • Molten globule state
  • Protein folding
  • Protein stability
  • Urea-induced denaturation

ASJC Scopus subject areas

  • Structural Biology
  • Biochemistry
  • Molecular Biology
  • Economics and Econometrics
  • General Energy


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