@article {99931, title = {Microsecond dynamics during the binding-induced folding of an intrinsically disordered protein.}, journal = {J Mol Biol}, year = {2021}, month = {2021 Sep 16}, pages = {167254}, abstract = {

Tau is an intrinsically disordered protein implicated in many neurodegenerative diseases. The repeat domain fragment of tau, tau-K18, is known to undergo a disorder to order transition in the presence of lipid micelles and vesicles, in which helices form in each of the repeat domains. Here, the mechanism of helical structure formation, induced by a phospholipid mimetic, sodium dodecyl sulfate (SDS) at sub-micellar concentrations, has been studied using multiple biophysical probes. A study of the conformational dynamics of the disordered state, using photoinduced electron transfer coupled to fluorescence correlation spectroscopy (PET-FCS) has indicated the presence of an intermediate state, I, in equilibrium with the unfolded state, U. The cooperative binding of the ligand (L), SDS, to I has been shown to induce the formation of a compact, helical intermediate (IL) within the dead time (\~{}37 {\textmu}s) of a continuous flow mixer. Quantitative analysis of the PET-FCS data and the ensemble microsecond kinetic data, suggests that the mechanism of induction of helical structure can be described by a U <-> I <-> IL <-> FL mechanism, in which the final helical state, FL, forms from IL with a time constant of 50-200 {\textmu}s. Finally, it has been shown that the helical conformation is an aggregation-competent state that can directly form amyloid fibrils.

}, issn = {1089-8638}, doi = {10.1016/j.jmb.2021.167254}, author = {Sen, Sreemantee and Kumar, Harish and Udgaonkar, Jayant B} } @article {70226, title = {Binding-induced folding under unfolding conditions: Switching between induced fit and conformational selection mechanisms.}, journal = {J Biol Chem}, volume = {294}, year = {2019}, month = {2019 Nov 08}, pages = {16942-16952}, abstract = {

The chemistry of protein-ligand binding is the basis of virtually every biological process. Ligand binding can be essential for a protein to function in the cell by stabilizing or altering the conformation of a protein, particularly for partially or completely unstructured proteins. However, the mechanisms by which ligand binding impacts disordered proteins or influences the role of disorder in protein folding is not clear. To gain insight into this question, the mechanism of folding induced by the binding of a Pro-rich peptide ligand to the SH3 domain of phosphatidylinositol 3-kinase unfolded in the presence of urea has been studied using kinetic methods. Under strongly denaturing conditions, folding was found to follow a conformational selection (CS) mechanism. However, under mildly denaturing conditions, a ligand concentration-dependent switch in the mechanism was observed. The folding mechanism switched from being predominantly a CS mechanism at low ligand concentrations to being predominantly an induced fit (IF) mechanism at high ligand concentrations. The switch in the mechanism manifests itself as an increase in the reaction flux along the IF pathway at high ligand concentrations. The results indicate that, in the case of intrinsically disordered proteins too, the folding mechanism is determined by the concentration of the ligand that induces structure formation.

}, issn = {1083-351X}, doi = {10.1074/jbc.RA119.009742}, author = {Sen, Sreemantee and Udgaonkar, Jayant B} } @article {56657, title = {Microsecond sub-domain motions and the folding and misfolding of the mouse prion protein.}, journal = {Elife}, volume = {8}, year = {2019}, month = {2019 Apr 26}, abstract = {

Protein aggregation appear to originate from partially unfolded conformations that are sampled through stochastic fluctuations of the native protein. It has been a challenge to characterize these fluctuations, under native like conditions. Here, the conformational dynamics of the full-length (23-231) mouse prion protein were studied under native conditions, using photoinduced electron transfer coupled to fluorescence correlation spectroscopy (PET-FCS). The slowest fluctuations could be associated with the folding of the unfolded state to an intermediate state, by the use of microsecond mixing experiments. The two faster fluctuations observed by PET-FCS, could be attributed to fluctuations within the native state ensemble. The addition of salt, which is known to initiate the aggregation of the protein, resulted in an enhancement in the time scale of fluctuations in the core of the protein. The results indicate the importance of native state dynamics in initiating the aggregation of proteins.

}, issn = {2050-084X}, doi = {10.7554/eLife.44766}, author = {Goluguri, Rama Reddy and Sen, Sreemantee and Udgaonkar, Jayant} } @article {20329, title = {A Dry Transition State More Compact Than the Native State Is Stabilized by Non-Native Interactions during the Unfolding of a Small Protein.}, journal = {Biochemistry}, year = {2017}, month = {2017 Jul 11}, abstract = {

Defining the role of non-native interactions in directing the course of protein folding or unfolding reactions has been a difficult challenge. In particular, the extent to which such interactions play a productive role by stabilizing the structures of transition states (TSs) found on the folding and unfolding pathways of proteins is not known. On the contrary, it is thought that the TSs are expanded forms of the N state stabilized by native interactions, and it is not known whether non-native interactions can modulate TS structure. In this study of the unfolding of the SH3 domain of PI3 kinase using a microsecond mixing methodology, partial non-native structure formation is shown to occur initially during unfolding. The TS of this partial "folding during unfolding" reaction is more compact than the N state: the apparent rate constant of Trp53 burial during this reaction decreases with an increase in denaturant concentration. Kinetic studies of the unfolding of mutant variants suggest that the unusually compact TS is stabilized by interactions not present in N and that these non-native interactions are hydrophobic in nature. It was determined that mutation could be used to tune the degree of compaction in the TS.

}, issn = {1520-4995}, doi = {10.1021/acs.biochem.7b00388}, author = {Sen, Sreemantee and Goluguri, Rama Reddy and Udgaonkar, Jayant B} }