Evidence for domain motion in proteins affecting global diffusion properties: A nuclear magnetic resonance study

The rotational diffusion of proteins is an important hydrodynamic property. Compact protein structures were found previously to exhibit hydration layer viscosity, η_loc, higher than the viscosity of bulk water, η. This implies an apparent activation energy for rotational diffusion higher than the activation energy of water viscosity, Eη = 15.4 ± 0.3 kJ/mol. In this study we examine η_loc of internally mobile proteins using ¹⁵N spin relaxation methods. We also examine the activation enthalpy, ΔH^#, and activation entropy, ΔS^#, for rotational diffusion. Of particular relevance are internally mobile ligand-free forms and compact ligand-bound forms of multidomain proteins. Adenylate kinase (AKeco) and Ca²⁺-calmodulin (Ca²⁺-CaM) are typical examples. For AKeco (Ca²⁺-CaM) we find that ΔH^# is 14.5 ± 0.5 (15.7 ± 0.4) kJ/mol. For the complex of AKeco with the inhibitor AP₅A (the complex of Ca²⁺-CaM with the peptide smMLCKp), we find that ΔH^# is 18.1 ± 0.7 (18.2 ± 0.5) kJ/mol. The internally mobile outer surface protein A has ΔH ^# = 12.6 ± 0.8 kJ/mol, and the compact protein Staphylococcal nuclease has ΔH^# = 18.8 ± 0.6 kJ/mol. For the internally mobile and compact proteins studied, <|ΔS^#|> equals 62 ± 7 J/(mol K) and 44 ± 5 J/(mol K), respectively. The fact is that η_loc > η (ΔH^# > Eη) for compact proteins was ascribed previously to electrostatic interactions between surface sites and water rigidifying the hydration layer. We find herein that obliteration of these interactions by domain motion leads to η^loc ∼ η, ΔH^# ∼ Eη, and large activation entropy for internally mobile protein structures.