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Crystal structure of the Cys-NO modified YopH tyrosine phosphatase

DOI: 10.1016/j.bbapap.2022.140754 DOI Help

Authors: Ruth F. Rocha (Universidade Federal de Santa Catarina) , Priscila G. A. Martins (Universidade Federal de Santa Catarina) , Humberto D'Muniz Pereira (Universidade de São Paulo) , Jose Brandao-Neto (Diamond Light Source) , Otavio Henrique Thiemann (University of São Paulo; Federal University of São Carlos) , Hernán Terenzi (Universidade Federal de Santa Catarina) , Angela C. O. Menegatti (Federal University of Paraib)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Biochimica Et Biophysica Acta (bba) - Proteins And Proteomics , VOL 1870

State: Published (Approved)
Published: March 2022
Diamond Proposal Number(s): 25296

Abstract: Protein tyrosine phosphatases (PTPs) are key virulence factors in pathogenic bacteria, consequently, they have become important targets for new approaches against these pathogens, especially in the fight against antibiotic resistance. Among these targets of interest YopH (Yersinia outer protein H) from virulent species of Yersinia is an example. PTPs can be reversibly inhibited by nitric oxide (NO) since the oxidative modification of cysteine residues may influence the protein structure and catalytic activity. We therefore investigated the effects of NO on the structure and enzymatic activity of Yersinia enterocolitica YopH in vitro. Through phosphatase activity assays, we observe that in the presence of NO YopH activity was inhibited by 50%, and that this oxidative modification is partially reversible in the presence of DTT. Furthermore, YopH S-nitrosylation was clearly confirmed by a biotin switch assay, high resolution mass spectrometry (MS) and X-ray crystallography approaches. The crystal structure confirmed the S-nitrosylation of the catalytic cysteine residue, Cys403, while the MS data provide evidence that Cys221 and Cys234 might also be modified by NO. Interestingly, circular dichroism spectroscopy shows that the S-nitrosylation affects secondary structure of wild type YopH, though to a lesser extent on the catalytic cysteine to serine YopH mutant. The data obtained demonstrate that S-nitrosylation inhibits the catalytic activity of YopH, with effects beyond the catalytic cysteine. These findings are helpful for designing effective YopH inhibitors and potential therapeutic strategies to fight this pathogen or others that use similar mechanisms to interfere in the signal transduction pathways of their hosts.

Journal Keywords: PTP; YopH; S-nitrosylation; YopH structure; Phosphatase inhibition; Nitric oxide

Diamond Keywords: Bacteria

Subject Areas: Biology and Bio-materials, Medicine, Chemistry

Instruments: I04-1-Macromolecular Crystallography (fixed wavelength)

Added On: 10/01/2022 09:58

Discipline Tags:

Pathogens Infectious Diseases Health & Wellbeing Biochemistry Chemistry Structural biology Drug Discovery Life Sciences & Biotech

Technical Tags:

Diffraction Macromolecular Crystallography (MX)