Structural and Dynamical Properties of a Partially Unfolded Fe₄S₄ Protein:  Role of the Cofactor in Protein Folding

Abstract

Heteronuclear multidimensional NMR spectroscopy was used to investigate in detail the structural and dynamical properties of a partially unfolded intermediate of the reduced high-potential iron-sulfur protein (HiPIP) from Chromatium vinosum present in 4 M guanidinium chloride solution. After an extensive assignment of 15N and 1H resonances, NOE data, proton longitudinal relaxation times, and 3JHNHalpha coupling constants as well as 15N relaxation parameters (T1, T2, T1rho, and 1H-15N NOE) were obtained and used to build a structural model of the intermediate. The Fe4S4 cluster of the HiPIP plays a decisive role in determining the resulting structure, which is random in the N-terminal half of the protein and partially organized in the loops between the cysteines bound to the cluster. Consistent with the structural data, the backbone mobility is typical of folded proteins in the regions where there are elements of structure and increases with the structural indetermination.

Functional Tat Transport of Unstructured, Small, Hydrophilic Proteins

“…We chose HiPIP, which is the smallest established Tat substrate so far. HiPIP requires the assembly of a [4Fe-4S] 2ϩ/3ϩ cofactor for folding, since the cluster assembly induces folding, and the loss of the cluster results in unstructured protein (36,37). In addition, HiPIP has no large hydrophobic core region.…”

1 H NMRD profiles of diamagnetic proteins: a model‐free analysis†

“…With both equations the fits are almost equally good, and often indistinguishable in terms of theoretical curves. The solid lines in Figs 1-4 represent the model-free fits performed with Eqns (3) and (6). The corresponding best fit parameters are reported in Table 1: the Cole-Cole values are reported next to the column of the Lorentzian 0 values.…”

“…In Eqns (1) and (3), the spectral density function takes the form 32 J ω D Equation (6) has been shown to be valid for 'any stationary Markov process obeying detailed balance, which includes virtually all dynamic models of interest,' 32 and is used to fit the experimental data in the present work. The number of terms N that can be included in Eqn (6) without unreasonably increasing the number of parameters 2N C 2 can be determined by a merit function analysis. 32 For the present data a sum of three Lorentzians turned out to be the right choice in all cases.…”

Bioinorganic chemistry in the postgenomic era

“…It can be anticipated, however, that this picture is not entirely true, and that these systems may actually be active in signaling their need for metal ions (42), thus playing a more relevant role in the broader picture of the cellular processes. Finally, metal ions play a prominent role in protein folding͞un-folding (43,44), as well as neurodegenerative diseases (45,46).…”