Biology subjects

Vasa, S. K.

Publications and source records attributed to Vasa, S. K..

4 recordsLinked to original sources

bioRxiv2024

Allostery at a protein-protein interface harboring an intermolecular dynamic network

Motional properties of individual amino acids in proteins are strongly modulated by their specific surrounding. The dynamics of tightly interacting residues can form intramolecular dynamic networks, which influence various features of protein function and serve as an access point for their modulation within signaling cascades. However, the possible formation of intermolecular networks shared between natural or constructed interaction partners has escaped thorough experimental assessment. Here, using fast-MAS solid-state NMR spectroscopy, we contrast the absence of a cross-talk between different residues in an apo protein with a recoupling of s timescale dynamics effective via a mediating crystal-crystal contact. The data show that dynamic allostery is not necessarily restricted to motionally coupled elements within a single protein but can traverse molecular boundaries. Interrogation of intermolecular dynamic networks by the strategies proposed here may shed light on the mechanisms underlying allosteric modulation of protein function in biological, pharmacological, and biotechnological studies.

biophysics↗

bioRxiv2024

Protein deuteration via algal amino acids to overcome proton back-exchange for fast-MAS solid-state NMR of large proteins

With perdeuteration, a current standard for solid-state NMR spectroscopy, large proteins suffer from incomplete amide-proton back-exchange. Using a 72 kDa micro-crystalline protein, we show that deuteration exclusively via deuterated amino acids, largely suppressing sidechain protonation, provides spectral resolution comparable to perdeuterated preparations at intermediate spinning frequencies without proton back-exchange obstacles.

biochemistry↗

bioRxiv2022

Integrated assessment of structure and dynamics of solid proteins via accurate solid-state NMR distance information

Understanding macromolecular function, interactions and stability hinges on detailed assessment of conformational ensembles. For solid proteins, accurate elucidation of the spatial aspects of dynamics at physiological temperatures are limited by the qualitative character or low abundance of solid-state NMR internuclear distance information. Here, we demonstrate access to abundant proton-proton internuclear distances for integrated structural biology and chemistry with unprecedented accuracy. Apart from highest-resolution single-state structures, the exact distances enable molecular dynamics (MD) ensemble simulations orchestrated by a dense network of experimental inter-proton distance boundaries gathered in the context of their physical lattices. This direct embedding of experimental ensemble distances into MD will provide access to representative, atomic-level spatial details of conformational dynamics in supramolecular assemblies, crystalline and lipid-embedded proteins, and beyond.

biophysics↗

bioRxiv2021

Residue-specific insights into (2x)72 kDa tryptophan synthase obtained from fast-MAS 1H-detected solid-state NMR

Solid-state NMR has emerged as a potent technique in structural biology, suitable for the study of fibrillar, micro-crystalline, and membrane proteins. Recent developments in fast-magic-angle-spinning and proton-detected methods have enabled detailed insights into structure and dynamics, but molecular-weight limitations for the asymmetric part of target proteins have remained at ~30-40 kDa. Here we employ solid-state NMR for atom-specific characterization of the 72 kDa (asymmetric unit) microcrystalline protein tryptophan synthase, an important target in pharmacology and biotechnology, chemical-shift assignments of which we obtain via higher-dimensionality, 4D and 5D solid-state NMR experiments. The assignments for the first time provide comprehensive data for assessment of side chain chemical properties involved in the catalytic turnover, and, in conjunction with first-principles calculations, precise determination of thermodynamic and kinetic parameters is demonstrated for the essential acid-base catalytic residue {beta}K87. The insights provided by this study expand by nearly a factor of two the size limitations widely accepted for NMR today, demonstrating the applicability of solid-state NMR to systems that have been thought to be out of reach due to their complexity.

biochemistry↗