Spectroscopic and MD simulation studies on unfolding processes of mitochondrial carbonic anhydrase VA induced by urea

Danish Idrees, Amresh Prakash, Md Anzarul Haque, Asimul Islam, Faizan Ahmad, Md Imtaiyaz Hassan

Research output: Contribution to journalArticlepeer-review

27 Scopus citations


Carbonic anhydrase VA (CAVA) is primarily expressed in the mitochondria and involved in numerous physiological processes including lipogenesis, insulin secretion from pancreatic cells, ureagenesis, gluconeogenesis and neuronal transmission. To understand the biophysical properties of CAVA, we carried out a reversible urea-induced isothermal denaturation at pH 7.0 and 25°C. Spectroscopic probes, [θ]222 (mean residue ellipticity at 222 nm), F344 (Trp-fluorescence emission intensity at 344 nm) and Δε280 (difference absorption at 280 nm) were used to monitor the effect of urea on the structure and stability of CAVA. The urea-induced reversible denaturation curves were used to estimate ΔG0D , Gibbs free energy in the absence of urea; Cm, the mid-point of the denaturation curve, i.e. molar urea concentration ([urea]) at which ΔGD = 0; and m, the slope (=∂ΔGD/∂[urea]). Coincidence of normalized transition curves of all optical properties suggests that unfolding/refolding of CAVA is a two-state process. We further performed 40 ns molecular dynamics simulation of CAVA to see the dynamics at different urea concentrations. An excellent agreement was observed between in silico and in vitro studies.

Original languageEnglish (US)
Pages (from-to)1987-1997
Number of pages11
JournalJournal of Biomolecular Structure and Dynamics
Issue number9
StatePublished - Sep 1 2016
Externally publishedYes


  • carbonic anhydrase VA
  • isothermal denaturation
  • molecular dynamics simulation
  • protein folding
  • protein stability
  • two-state unfolding
  • urea-induced denaturation

ASJC Scopus subject areas

  • Structural Biology
  • Molecular Biology


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