Temperature-resolved Cryo-EM Uncovers Structural Bases of Temperature-dependent Enzyme Functions.

By Chin-Yu Chen, Yuan-Chih Chang, B. Lin, Chun-Hsiang Huang, M. Tsai. 2019

28 citations recorded by the source corpus.

Protein functions are temperature-dependent, but protein structures are usually solved at a single, often low, temperature due to limitations in the conditions of crystal growth or protein vitrification. Here we demonstrate the feasibility of solving cryo-EM structures of proteins vitrified at high temperatures, solve 12 structures of an archaeal ketol-acid reductoisomerase (KARI) vitrified at 4-70 °C, and show that structures of both the Mg2+-form (KARI:2Mg2+) and its ternary complex (KARI:2Mg2+:NADH:inhibitor) are temperature-dependent in correlation with the temperature dependence of enzyme activity. Furthermore, structural analyses led to dissection of the induced-fit mechanism into ligand-induced and temperature-induced effects, and to capture temperature-resolved intermediates of the temperature-induced conformational change. The results also suggest that it is preferable to solve cryo-EM structures of protein complexes at functional temperatures. These studies should greatly expand the landscapes of protein structure-function relationship, and enhance the mechanistic analysis of enzymatic functions.

Subfields: Materials Chemistry, Molecular Biology, Structural Biology

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Citations in the corpus

State-specific protein–ligand complex structure prediction with a multiscale deep generative model (2022, positive): “5f) whose catalytic mechanism was recently characterized 50 , by comparing the predicted holo and apo ensembles, we find that NeuralPLexer accurately captures the closure motion of the N-subdomain upon cofactor and inhibitor binding.”
Advances in methods for atomic resolution macromolecular structure determination (2020, background): “An emerging frontier in cryo-EM data interpretation is the analysis of energetic landscapes and the dissection of structural heterogeneity 94 .”