Cold Shock Domain of the Human Y-Box Protein YB-1. Backbone Dynamics and Equilibrium between the Native State and a Partially Unfolded State

Abstract

The three-dimensional structure of the central cold shock domain (CSD) of the human Y-box protein (YB-1 CSD) is virtually identical to those available for the bacterial cold shock proteins (Csp's). We have further characterized YB-1 CSD by studying its dynamics by nuclear magnetic resonance. The observed structural similarity is reflected in the backbone dynamics, which for YB-1 CSD is very similar to that of the Escherichia coli protein CspA. The rotational correlation time of YB-1 CSD shows that it is a monomer. This indicates that the dimerization observed for the YB-1 protein is not caused by its CSD, but involves other parts of this protein. The YB-1 CSD is only marginally stable as are the mesophilic bacterial Csp's. In contrast to the rapid two-state folding of the bacterial Csp's, the formation of the native form of YB-1 CSD is slow and at least a three-state process. The NMR experiments revealed the presence of a second state of YB-1 CSD in equilibrium with the native form. The exchange rates from and to the folded state are in the order of 0.2 and 0.5 s(-1), respectively. Relaxation experiments indicated that the second state is a highly flexible, partly structured molecule.

Structural basis of DNA binding to human YB-1 cold shock domain regulated by phosphorylation

“…One set is characteristic of disordered (unfolded) proteins with 1 H chemical shifts in a narrow range of ∼7.9–8.5 ppm, while the other is characteristic of folded proteins. This is consistent with the previous study ( 45 ). The result shows that this CSD construct exists in two conformations, one folded and one unfolded or disordered, which undergo slow conformational exchange in the NMR time regime.…”

“…Peak assignments of CSDex are labeled. For CSD (aa 51–129), the assignments were retrieved from the published data ( 45 ) and the residues from the unfolded form are indicated by appending ‘U’ in front of residue numbers. The peaks from the sidechains of W, Q and N are labeled by appending “s" after the residue numbers.…”

Y-Box Binding Proteins in mRNP Assembly, Translation, and Stability Control

“…The YB-1 cold shock domain consists of five anti-parallel β-strands forming a β-barrel. The NMR analysis showed low stability of CSD that remains only 70% native at 25 °C [14]. This fact is also supported by microcalorimetry data.…”

Thermodynamic characterization of the interaction between the human Y-box binding protein YB-1 and nucleic acids

“…The CD spectrum of YB-1(Full) is similar to that of FRGY2 (Y-box protein from Xenopus laevis) 19 and YB-1 from Guryanov's group reported previously. 20 Although it has been reported that the CSD of YB-1 has an antiparallel b-barrel structure in only 95% H 2 O and 5% D 2 O solution, 21 our CD data showed that the domain had mainly random coils under the buffer conditions. The NMR relaxation measurements have demonstrated reportedly that the CSD exhibits an equilibrium between 70% folded and 30% unfolded states.…”