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A disulfide driven domain swap switches off the activity of Shigella IpaH9.8 E3 ligase |
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| Authors: | Arefeh Seyedarabi James A Sullivan Richard W Pickersgill |
| Institution: | a School of Biological and Chemical Sciences, Queen Mary University of London, Mile End Road, London E1 4NS, United Kingdom b Department of Microbiology and Immunology and Department of Infectious Disease Control, International Research Center for Infectious Disease, Institute of Medical Science, University of Tokyo, 4-6-1, Shirokanedai, Minato-ku, Tokyo 108-8639, Japan |
| Abstract: | We show that the monomeric form of Shigella IpaH9.8 E3 ligase catalyses the ubiquitination of human U2AF35 in vitro, providing a molecular mechanism for the observed in vivo effect. We further discover that under non-reducing conditions IpaH9.8 undergoes a domain swap driven by the formation of a disulfide bridge involving the catalytic cysteine and that this dimer is unable to catalyse the ubiquitination of U2AF35. The crystal structure of the domain-swapped dimer is presented. The redox inactivation of IpaH9.8 could be a mechanism of regulating the activity of the IpaH9.8 E3 ligase in response to cell damage so that the host cell in which the bacteria resides is maintained in a benign state suitable for bacterial survival.Structured summaryMINT-7993779: ipaH9.8 (uniprotkb:Q8VSC3) and ipaH9.8 (uniprotkb:Q8VSC3) bind (MI:0408) by X-ray crystallography (MI:0114) MINT-7993812: ipaH9.8 (uniprotkb:Q8VSC3) and ipaH9.8 (uniprotkb:Q8VSC3) bind (MI:0407) by affinity chromatography technology (MI:0004) MINT-7993790: ipaH9.8 (uniprotkb:Q8VSC3) and ipaH9.8 (uniprotkb:Q8VSC3) bind (MI:0407) by blue native page (MI:0276) |
| Keywords: | AP alkaline phosphotase DTT dithiotheritol GST glutathione S-transferase His-tag hexahistidine-tag HRP horse raddish peroxidase LRR leucine rich repeat RT room temperature Me methylated MMC methylmercury chloride mRNA messenger ribonucleic acid ORF open reading frame PCR polymerase chain reaction PDB protein data bank SDS sodium dodecyl sulphate TCEP tris (2-carboxyethyl) phosphine hydrochloride WT wild type |
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