S-Nitrosylation Induces Structural and Dynamical Changes in a Rhodanese Family Protein

•S-nitrosylation is a common protein modification, but its impact on structure and dynamics is not well understood.•The rhodanese domain of YgaP has a rhodanese-typical conformation of the active loop with the catalytic cysteine sandwiched between the N termini of α4 and α5 helices.•S-nitrosylation and S-sulfhydration are competing processes in YgaP and both are present when YgaP is overexpressed in E. coli.•S-sulfhydration destabilizes helix α4, but under nitrosative stress, the balance shifts toward S-nitrosylation, which induces the slow dynamics of helix α5 while stabilizing α4.

S-Nitrosylation is well established as an important post-translational regulator in protein function and signaling. However, relatively little is known about its structural and dynamical consequences. We have investigated the effects of S-nitrosylation on the rhodanese domain of the Escherichia coli integral membrane protein YgaP by NMR, X-ray crystallography, and mass spectrometry. The results show that the active cysteine in the rhodanese domain of YgaP is subjected to two competing modifications: S-nitrosylation and S-sulfhydration, which are naturally occurring in vivo. It has been observed that in addition to inhibition of the sulfur transfer activity, S-nitrosylation of the active site residue Cys63 causes an increase in slow motion and a displacement of helix 5 due to a weakening of the interaction between the active site and the helix dipole. These findings provide an example of how nitrosative stress can exert action at the atomic level.

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