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Allosteric cooperativity plays a critical role in the function of biological complexes. Methyl-TROSY NMR spectroscopy is a powerful technique to probe the structure and dynamics of large molecular complexes and is well suited to investigating allostery and cooperativity in such systems. In this chapter, we discuss the general concepts and models of cooperativity and introduce strategies to probe cooperativity in high-molecular-weight complexes by methyl-TROSY NMR. We include an overview of the methyl-TROSY technique, focusing on the practical aspects of isotope labeling of the methyl groups and current approaches for methyl resonance assignment. We present a few case studies that exemplify the unique advantages of the methyl-TROSY approach in providing insights into cooperativity and allostery in functionally important molecular machines.

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