title: Characterization of folding intermediates during urea-induced denaturation of human carbonic anhydrase II.
abstract: Knowledge of folding/unfolding pathway is fundamental basis to study protein structure and stability. Human carbonic anhydrase II (HCAII) is a ∼29kDa, 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 XIand XIIpopulated around 2 and 4M urea, respectively. The far-UV CD was used to characterize the intermediate states (XIand 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 XIstate has characteristics of molten globule state while XIIstate 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.
authors: Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, Jamia Nagar, New Delhi 110025, India.
authors: Wahiduzzaman
authors: Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, Jamia Nagar, New Delhi 110025, India.
authors: Mohammad Aasif
authors: MA
authors: Dar
authors: Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, Jamia Nagar, New Delhi 110025, India.
authors: Md Anzarul
authors: MA
authors: Haque
authors: Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, Jamia Nagar, New Delhi 110025, India.
authors: Danish
authors: D
authors: Idrees
authors: Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, Jamia Nagar, New Delhi 110025, India.
authors: Md Imtaiyaz
authors: MI
authors: Hassan
authors: Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, Jamia Nagar, New Delhi 110025, India. Electronic address: aislam@jmi.ac.in.
authors: Asimul
authors: A
authors: Islam
authors: Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, Jamia Nagar, New Delhi 110025, India.
authors: Faizan
authors: F
authors: Ahmad
mesh_terms: D024402:Carbonic Anhydrase II / Q000737:chemistry*; D006801:Humans; D057446:Hydrodynamics; D057927:Hydrophobic and Hydrophilic Interactions; D011489:Protein Denaturation / Q000187:drug effects*; D017434:Protein Structure, Tertiary / Q000187:drug effects; D014508:Urea / Q000494:pharmacology*
chemical_list: D014508:Urea; D024402:Carbonic Anhydrase II
keywords: Human carbonic anhydrase II; Molten globule state; Protein folding; Protein stability; Urea-induced denaturation