Equilibrium folding dynamics of meACP in water, heavy water, and low concentration of urea

Abstract

Many proteins fold in apparent two-state behavior, as partially folded intermediates only transiently accumulate and easily escape detection. Besides a native form and a mainly unfolded form, we captured a partially unfolded form of an acyl carrier protein from Micromonospora echinospora (meACP) in the folding/unfolding equilibrium using chemical exchange saturation transfer NMR experiments. The C-terminal region of the partially unfolded form is mainly folded and the N-terminal is unfolded. Furthermore, to understand how the folding process of meACP is influenced by solvent environments, we compared the folding dynamics of meACP in D2O, H2O and low concentration of urea. As the environment becomes more denaturing from D2O to H2O and then to urea, the unfolded state becomes increasingly populated, and the folding rate decreases. Adding a small amount of urea, which does not change solvent viscosity, has little effects on the unfolding rates, while changing H2O to D2O reduces the unfolding rates possibly due to the increase of solvent viscosity. The quantified solvent effects on the protein folding Gibbs energy and activation energy suggest that the transition state of folding may have a similar structure to the native state of the protein.

Mechanism and quantitative assessment of saturation transfer for water‐based detection of the aliphatic protons in carbohydrate polymers

“…Assuming the slow tumbling limit, then and (with f equal to k k ), Equation (7) resembles the analytical solution for the proton transfer enhancement factor in the 2-pool CEST model, 24 however, now with the NOE transfer as the determining rate instead of k . Equations (4) to (7) indicate that the relative values of the NOE rate and the hydroxyl proton exchange rate are important in determining the glycoNOE signals at different pH values. When k ≫| |, the rNOE signal in the Z-spectrum is expected to be pH-independent, which will be expected for glycogen under physiological conditions.…”

“…In the CEST MRI method, a RF pulse is applied to saturate the protons of targeted molecules, and the change in water signal due to saturation transfer is quantified as a function of saturation frequency in the Z-spectrum. [1][2][3][4][5] Magnetization transfer (MT) between protons may occur via several mechanisms: 1) through-space cross-relaxation (or nuclear Overhauser effect [NOE]); or 2) exchange processes, including conformational exchange, 6,7 ligand binding, 8,9 protein hydration, 10 and labile (hydroxyl, amide, or amine) proton exchange with water. 2 Due to the abundance of proton MT processes in vivo, multiple signals appear in the output Z-spectrum.…”

Structural Characterization of an ACP from Thermotoga maritima: Insights into Hyperthermal Adaptation

“…ACPs consist of four helical bundles connected by three loop regions, forming hydrophobic cavities to accommodate the growing acyl chains. ACPs are believed to be dynamic proteins, and their flexibilities are essential for their functions [10,[24][25][26][27][28]. Vh-ACP.…”

Salt Dependence Conformational Stability of the Dimeric SAM Domain of MAPKKK Ste11 from Budding Yeast: A Native-State H/D Exchange NMR Study

“…[29][30][31] It reveals the potential metastable, partially unfolded intermediates, their interaction (exchange) behavior with the solvent, as well as the denaturant, that occur in the steps leading up to the completely unfolded state. [32][33][34] Such transitions are mostly cooperative and may involve a sequential unfolding of the structural units in tandem with their underlying counterparts. 35,36 In the present study, we elucidate the effect of monovalent salt in stabilizing the hydrophobic interfaces of the Ste11 SAM domain under urea-induced mild denaturing conditions.…”

“…As per the Boltzmann distribution, under native-state conditions, any protein molecule has a small population of transient state intermediates occupying a higher than ground state energy level, called the partially unfolded forms (PUFs) and even under mildly denaturing condition protein molecules will cycle through these globally unfolded states. , The prolonged hydrogen exchange (HX) behavior thus can be utilized to study the global protein unfolding pathway; the identification and study of these intermediates, therefore, pave the way for elucidating the mechanism of protein cooperativity as well protein folding and unfolding. − The hydrogen exchange study of native protein under mild denaturing conditions is realized through different procedures. − It reveals the potential metastable, partially unfolded intermediates, their interaction (exchange) behavior with the solvent, as well as the denaturant, that occur in the steps leading up to the completely unfolded state. − Such transitions are mostly cooperative and may involve a sequential unfolding of the structural units in tandem with their underlying counterparts. , …”