Delineating the conformational dynamics of intermediate structures on the unfolding pathway of β-lactoglobulin in...

The funnel shaped energy landscape model of the protein folding suggests that progression of folding proceeds through multiple pathways, having the multiple intermediates which leads to multidimensional free-energy surface. Herein, we applied all-atom MD simulation to conduct a comparative study on the structure of β-lactoglobulin (β-LgA) in aqueous mixture of 8 M urea and 8 M dimethyl sulfoxide (DMSO), at different temperatures. The cumulative results of multiple simulations suggest a common unfolding pathway of β-LgA, occurred through the stable and meta-stable intermediates (I), in both urea and DMSO. However, the free-energy landscape (FEL) analyses show that the structural transitions of I-states are energetically different. In urea, FEL shows distinct ensemble of intermediates, I1 and I2, separated by the energy barrier of ∼3.0 kcal mol^-1. Similarly, we find the population of two distinct I1 and I2 states in DMSO, however, the I1 appeared transiently around ∼30-35 ns and is short-lived. But, the I2 ensemble is observed structurally compact and long-lived (∼50-150 ns) as compared to unfolding in urea. Furthermore, the I1 and I2 are separated through a high energy barrier of ∼6.0 kcal mol^-1. Thus, our results provide the structural insights of intermediates which essentially bear the signature of a different unfolding pathway of β-LgA in urea and DMSO.Abbreviationsβ-LgAβ-lactoglobulinDMSOdimethyl sulfoxideFELfree-energy landscapeGdmClguanidinium chlorideIintermediate stateMGmolten globule statePMEparticle mesh EwaldQfraction of native contactsRMSDroot mean square deviationRMSFroot mean square fluctuationR_gradius of gyrationSASAsolvent Accessible Surface AreascSASAthe side chain SASATrptryptophanCommunicated by Ramaswamy H. Sarma.