PPAD

P. gingivalis PAD (PPAD) ‐ Citrullination is a post‐translational modification ofhigher organisms that deiminates arginines in proteins and peptides. It occurs in physiological processes but also pathologies such as multiple sclerosis, fibrosis, Alzheimer’s disease and rheumatoid arthritis (RA). The reaction is catalyzed by peptidylarginine deiminases (PADs), which are found in vertebrates but not in lower organisms. Uniquely among microbes to date, Porphyromonas gingivalis also produces a secreted PAD (called PPAD), which protectsP. gingivalisduring acidic cleansing in the mouth through ammonia generated during host and endogenous protein citrullination. PPAD does not require calcium for catalysis and is genetically unrelated with animal PADs and, like the latter and cysteine peptidases, its main catalytic residue is a cysteine (C351 in PPAD). Human PADs citrullinates arginines within polypeptide chains but not at their termini, i.e. they are efficient endodeiminases but poor exodeiminases. In contrast, PPAD citrullinates C‐terminal arginines commonly generated by the proteolytic action of gingipain R (Rgps), further enhanced by the surface co‐localization of Rgps and PPAD. It comprises a flat cylindrical catalytic domain with five‐fold α/β‐propeller architecture and a C‐terminal immunoglobulin‐like domain. Catalysis is based on a cysteine‐histidine‐asparagine triad, which is shared with human PAD1‐PAD4 and other guanidino‐group modifying enzymes.

Peptidyl‐arginine deiminase from Porphyromonas gingivalis

Uniprot Q9RQJ2 / GeneBank locus name: PG_1424 / PDB: 4YT9

Goulas T, Mizgalska D, Garcia‐Ferrer I, Kantyka T, Guevara T, Szmigielski B, Sroka A, Millán C, Usón I, Veillard F, Potempa B, Mydel P, Solà M, Potempa J, Gomis‐Rüth FX. (2015) Structure and mechanism of a bacterial host‐protein citrullinating virulence factor, Porphyromonas gingivalis peptidylarginine deiminase. Sci Rep. Jul 1;5:11969.

Purified from growth medium of genetically modified P. gingivalis expressing active short form of PPAD (catalytic domain + IgSF domain) with C‐terminal hexahistidine‐tag.

Aceton precipitation, DE‐52 cellulose ion‐exchange chromatography, Ni‐Sepharose affinity chromatography, MonoQ ion‐exchange chromatography, size exclusion chromatography

~46 kDa

30 μg / minimum 750 mU

batch specific (≥90% estimated by SDS‐PAGE)

20 mM Tris, pH 7.5

Dry ice

‐ 80°C. Avoid freeze/thaw cycles by aliquoting protein

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