Nanosecond-Timescale Dynamics and Conformational Heterogeneity in Human GCK Regulation and Disease
| Autor: | A. Carl Whittington, Shawn M. Sternisha, Gianluigi Veglia, Malcolm M. McCray, Juliana A. Martinez Fiesco, Brian G. Miller, Carol M. Porter, Timothy M. Logan, Peter J. Steinbach, Cristina Olivieri |
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| Rok vydání: | 2019 |
| Předmět: |
Magnetic Resonance Spectroscopy
Kinetics Allosteric regulation Population Biophysics Molecular Conformation Cooperativity medicine.disease_cause 03 medical and health sciences 0302 clinical medicine Glucokinase medicine Humans education 030304 developmental biology 0303 health sciences Mutation education.field_of_study Chemistry Nuclear magnetic resonance spectroscopy Articles Glucose binding 030217 neurology & neurosurgery |
| Zdroj: | Biophys J |
| ISSN: | 1542-0086 |
| Popis: | Human glucokinase (GCK) is the prototypic example of an emerging class of proteins with allosteric-like behavior that originates from intrinsic polypeptide dynamics. High-resolution NMR investigations of GCK have elucidated millisecond-timescale dynamics underlying allostery. In contrast, faster motions have remained underexplored, hindering the development of a comprehensive model of cooperativity. Here, we map nanosecond-timescale dynamics and structural heterogeneity in GCK using a combination of unnatural amino acid incorporation, time-resolved fluorescence, and 19F nuclear magnetic resonance spectroscopy. We find that a probe inserted within the enzyme’s intrinsically disordered loop samples multiple conformations in the unliganded state. Glucose binding and disease-associated mutations that suppress cooperativity alter the number and/or relative population of these states. Together, the nanosecond kinetics characterized here and the millisecond motions known to be essential for cooperativity provide a dynamical framework with which we address the origins of cooperativity and the mechanism of activated, hyperinsulinemia-associated, noncooperative variants. |
| Databáze: | OpenAIRE |
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