Exploring tryptophan dynamics in acid-induced molten globule state of bovine α-lactalbumin: a wavelength-selective fluorescence approach

Abstract

The relevance of partially ordered states of proteins (such as the molten globule state) in cellular processes is beginning to be understood. Bovine alpha-lactalbumin (BLA) assumes the molten globule state at acidic pH. We monitored the organization and dynamics of the functionally important tryptophan residues of BLA in native and molten globule states utilizing the wavelength-selective fluorescence approach and fluorescence quenching. Quenching of BLA tryptophan fluorescence using quenchers of varying polarity (acrylamide and trichloroethanol) reveals varying degrees of accessibility of tryptophan residues, characteristic of native and molten globule states. We observed red edge excitation shift (REES) of 6 nm for the tryptophans in native BLA. Interestingly, we show here that BLA tryptophans exhibit REES (3 nm) in the molten globule state. These results constitute one of the early reports of REES in the molten globule state of proteins. Taken together, our results indicate that tryptophan residues in BLA in native as well as molten globule states experience motionally restricted environment and that the regions surrounding at least some of the BLA tryptophans offer considerable restriction to the reorientational motion of the water dipoles around the excited-state tryptophans. These results are supported by wavelength-dependent changes in fluorescence anisotropy and lifetime for BLA tryptophans. These results could provide vital insight into the role of tryptophans in the function of BLA in its molten globule state in particular, and other partially ordered proteins in general.

Spectroscopic Studies of Interaction of Protein with Cerium Oxide Nanoparticles

“…2. The fluorescence decays of BSA were fitted with two exponentials, T1 = 6.50 ns and T2 = 2.46 ns and is consistent with the studies that lifetimes of tryptophan fluorescence are often multi exponential [28]. The longer and shorter lifetimes indicated that BSA contained two tryptophan residues that fluoresced in two different environments [29] and one of the tryptophan residues in the protein may be buried inside the hydrophobic interior of protein whereas the other tryptophan residue may be close to quencher [30].…”

Interaction of Copper Oxide Nanoparticles With Bovine Serum Albumin by Spectroscopic Studies

“…The decrease was more prominent for lifetime T = 2.46 ns and is consistent with the studies that lifetimes of trp fluorescence are often multiexponential [29]. The changes in a lifetime, give information about the local environment of the trp-residues [30].…”

The red edge excitation shift phenomenon can be used to unmask protein structural ensembles: implications for NEMO–ubiquitin interactions

“…There have been a few reports where emission arising from a single Trp residue gives a significant REES effect in the folded and molten globule-like states [13], while the REES effect disappears or is rather reduced in unfolded states [14,15]. The REES effect is, therefore, a unique probe for proteins with a high degree of molecular flexibility resulting in an ensemble of solvent environments around the Trp probe.…”