Biology subjects

Udgaonkar, J. B.

Publications and source records attributed to Udgaonkar, J. B..

3 recordsLinked to original sources

bioRxiv2026

Solvent Isotope Effect on the Stability of a Heterodimeric Protein

Protein stability arises from a fine balance between stabilizing forces such as hydrophobic interactions, hydrogen bonding, and ionic interactions, and destabilizing contributions from solvent exposure and electrostatics. Although hydrophobic burial is the dominant driving force for folding, intra-chain hydrogen bonds and ionic interactions modulate stability in context-dependent ways, with effects that vary depending on their location and environment within the protein. Most studies of protein stability have focused on perturbations induced by pH, solvent composition, or mutations in protonated water, leaving the influence of solvent isotopes relatively underexplored. Notably, despite stronger hydrogen bonding in D2O, proteins exhibit diverse stability responses upon transfer from H2O to D2O, suggesting that differential hydration of nonpolar groups plays a key role. Here, the solvent isotope effect on protein stability is investigated using double-chain monellin (dcMN), a {beta}-sheet-rich, two-chain protein with well-characterized folding behavior. By combining conventional equilibrium unfolding measurements with hydrogen-deuterium exchange mass spectrometry (HDX-MS), the stability of wild-type and a less hydrophobic mutant (C42A) dcMN was compared in H2O and D2O, revealing greater stabilization of the wild-type protein in D2O and highlighting the importance of hydrophobic interactions in governing isotope-dependent stability.

biophysics↗

bioRxiv2023

HDX-MS reveals concealed conformations of ISWI during different stages of nucleosome sliding

Nucleosome spacing across the genome is regulated by the adenosine 5-triphosphate (ATP)- dependent nucleosome sliding activity of Imitation Switch, ISWI. ISWI is believed to be auto-inhibited in the resting-state by the binding of its N-and C-terminal regulatory regions to its central ATPase-domain, attaining a "closed" conformation. To slide nucleosomes ISWI must i) transition to the state competent for nucleosome binding, ii) bind to nucleosome and iii) carry the ATP-dependent nucleosome sliding. The conformations attained by full-length ISWI (FL-ISWI) during the entire sliding process have remained inaccessible by the methods used so far. Using Hydrogen/Deuterium-exchange coupled to Mass-Spectrometry (HDX-MS), we monitored conformational dynamics of the Drosophila FL-ISWI at all the stages of sliding process. HDX-MS data show that in the resting state, ISWI samples an ensemble of conformations showing varying levels of deuterium uptake in many regions including N-and C-terminal regulatory regions, suggesting ISWI intrinsically samples relatively "open-states". In addition to substantiating previous nucleosome binding studies, HDX-MS reveals that during actual sliding-step, regions of ATPase-domain which bind to the nucleosomal DNA undergo major conformational change. The C-terminal HSS domain switches from the solvent protected stable state to a more dynamic state, implying several interactions established by ISWI with the nucleosome upon binding are relieved during sliding. In sum, this study provides mechanistic insights into how ISWI can switch from an auto-inhibited "closed-state" to an "open-state" competent for nucleosome binding, and reveals the conformation attained by ISWI during the actual nucleosome sliding step. We propose that, like ISWI, intrinsic dynamics may be involved in functioning of other Rec-like ATPase-domain containing protein families.

biochemistry↗

bioRxiv2020

α-Synuclein aggregation intermediates form fibril polymorphs with distinct prion-like properties

-Synuclein (-Syn) amyloid fibrils in synucleinopathies (such as Parkinsons disease (PD), multiple system atrophy (MSA)) are structurally and functionally different, reminiscent of prion-like strains. However, how a single protein can form different fibril polymorphs in various synucleinopathies is not known. Here, we demonstrate the structure-function relationship of two distinct -Syn fibril polymorphs, the pre-matured fibrils (PMF) and helix-matured fibrils (HMF) based on -Syn aggregation intermediates. These polymorphs not only display the structural differences, including their fibril core structure as demonstrated by solid-state nuclear magnetic resonance (NMR) spectroscopy and H/D-exchange coupled with mass spectrometry but also possess different cellular activities such as seeding, cellular internalization, and cell-to-cell transmission. The HMF with a compact core structure exhibits low seeding potency in cells but readily internalizes and transmits from one cell to another. Whereas the less structured PMF lacks the cell-to-cell transmission ability but induces abundant -Syn pathology and triggers the formation of aggresomes in cells. Overall, the study highlights how the conformational heterogeneity in the aggregation pathway may lead to fibril polymorphs with distinct prion-like behavior in PD.

neuroscience↗