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Autocitrullination and Changes in the Activity of Peptidylarginine Deiminase 3 Induced by High Ca(2+) Concentrations

[Image: see text] Peptidylarginine deiminases (PADs) are enzymes that catalyze the Ca(2+)-dependent conversion of arginine residues into proteins to citrulline residues. Five PAD isozymes have been identified in mammals. Several studies have shown that the active-site pockets of these isozymes are f...

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Autores principales: Sawata, Mizuki, Shima, Hiroki, Murayama, Kazutaka, Matsui, Toshitaka, Igarashi, Kazuhiko, Funabashi, Kazumasa, Ite, Kenji, Kizawa, Kenji, Takahara, Hidenari, Unno, Masaki
Formato: Online Artículo Texto
Lenguaje:English
Publicado: American Chemical Society 2022
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9386831/
https://www.ncbi.nlm.nih.gov/pubmed/35990454
http://dx.doi.org/10.1021/acsomega.2c02972
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author Sawata, Mizuki
Shima, Hiroki
Murayama, Kazutaka
Matsui, Toshitaka
Igarashi, Kazuhiko
Funabashi, Kazumasa
Ite, Kenji
Kizawa, Kenji
Takahara, Hidenari
Unno, Masaki
author_facet Sawata, Mizuki
Shima, Hiroki
Murayama, Kazutaka
Matsui, Toshitaka
Igarashi, Kazuhiko
Funabashi, Kazumasa
Ite, Kenji
Kizawa, Kenji
Takahara, Hidenari
Unno, Masaki
author_sort Sawata, Mizuki
collection PubMed
description [Image: see text] Peptidylarginine deiminases (PADs) are enzymes that catalyze the Ca(2+)-dependent conversion of arginine residues into proteins to citrulline residues. Five PAD isozymes have been identified in mammals. Several studies have shown that the active-site pockets of these isozymes are formed when Ca(2+) ions are properly bound. We previously characterized the structures of PAD3 in six states. Among these, we identified a “nonproductive” form of PAD3 in which the active site was disordered even though five Ca(2+) ions were bound. This strange structure was probably obtained as a result of either high Ca(2+) concentration (∼260 mM)-induced denaturation during the crystallization process or high Ca(2+)-concentration-induced autocitrullination. While autocitrullination has been reported in PAD2 and PAD4 for some time, only a single report on PAD3 has been published recently. In this study, we investigated whether PAD3 catalyzes the autocitrullination reaction and identified autocitrullination sites. In addition to the capacity of PAD3 for autocitrullination, the autocitrullination sites increased depending on the Ca(2+) concentration and reaction time. These findings suggest that some of the arginine residues in the “nonproductive” form of PAD3 would be autocitrullinated. Furthermore, most of the autocitrullinated sites in PAD3 were located near the substrate-binding site. Given the high Ca(2+) concentration in the crystallization condition, it is likely that Arg372 was citrullinated in the “nonproductive” PAD3 structure, the structure was slightly altered from the active form by citrulline residues, and probably inhibited Ca(2+)-ion binding at the proper position. Following Arg372 citrullination, PAD3 enters an inactive form; however, the Arg372-citrullinated PAD3 are considered minor components in autocitrullinated PAD3 (CitPAD3), and CitPAD3 does not significantly decrease the enzyme activity. Autocitrullination of PAD3 could not be confirmed at the low Ca(2+) concentrations seen in vivo. Future experiments using cells and animals are needed to verify the effect of Ca(2+) on the PAD3 structure and functions in vivo.
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spelling pubmed-93868312022-08-19 Autocitrullination and Changes in the Activity of Peptidylarginine Deiminase 3 Induced by High Ca(2+) Concentrations Sawata, Mizuki Shima, Hiroki Murayama, Kazutaka Matsui, Toshitaka Igarashi, Kazuhiko Funabashi, Kazumasa Ite, Kenji Kizawa, Kenji Takahara, Hidenari Unno, Masaki ACS Omega [Image: see text] Peptidylarginine deiminases (PADs) are enzymes that catalyze the Ca(2+)-dependent conversion of arginine residues into proteins to citrulline residues. Five PAD isozymes have been identified in mammals. Several studies have shown that the active-site pockets of these isozymes are formed when Ca(2+) ions are properly bound. We previously characterized the structures of PAD3 in six states. Among these, we identified a “nonproductive” form of PAD3 in which the active site was disordered even though five Ca(2+) ions were bound. This strange structure was probably obtained as a result of either high Ca(2+) concentration (∼260 mM)-induced denaturation during the crystallization process or high Ca(2+)-concentration-induced autocitrullination. While autocitrullination has been reported in PAD2 and PAD4 for some time, only a single report on PAD3 has been published recently. In this study, we investigated whether PAD3 catalyzes the autocitrullination reaction and identified autocitrullination sites. In addition to the capacity of PAD3 for autocitrullination, the autocitrullination sites increased depending on the Ca(2+) concentration and reaction time. These findings suggest that some of the arginine residues in the “nonproductive” form of PAD3 would be autocitrullinated. Furthermore, most of the autocitrullinated sites in PAD3 were located near the substrate-binding site. Given the high Ca(2+) concentration in the crystallization condition, it is likely that Arg372 was citrullinated in the “nonproductive” PAD3 structure, the structure was slightly altered from the active form by citrulline residues, and probably inhibited Ca(2+)-ion binding at the proper position. Following Arg372 citrullination, PAD3 enters an inactive form; however, the Arg372-citrullinated PAD3 are considered minor components in autocitrullinated PAD3 (CitPAD3), and CitPAD3 does not significantly decrease the enzyme activity. Autocitrullination of PAD3 could not be confirmed at the low Ca(2+) concentrations seen in vivo. Future experiments using cells and animals are needed to verify the effect of Ca(2+) on the PAD3 structure and functions in vivo. American Chemical Society 2022-08-08 /pmc/articles/PMC9386831/ /pubmed/35990454 http://dx.doi.org/10.1021/acsomega.2c02972 Text en © 2022 The Authors. Published by American Chemical Society https://creativecommons.org/licenses/by-nc-nd/4.0/Permits non-commercial access and re-use, provided that author attribution and integrity are maintained; but does not permit creation of adaptations or other derivative works (https://creativecommons.org/licenses/by-nc-nd/4.0/).
spellingShingle Sawata, Mizuki
Shima, Hiroki
Murayama, Kazutaka
Matsui, Toshitaka
Igarashi, Kazuhiko
Funabashi, Kazumasa
Ite, Kenji
Kizawa, Kenji
Takahara, Hidenari
Unno, Masaki
Autocitrullination and Changes in the Activity of Peptidylarginine Deiminase 3 Induced by High Ca(2+) Concentrations
title Autocitrullination and Changes in the Activity of Peptidylarginine Deiminase 3 Induced by High Ca(2+) Concentrations
title_full Autocitrullination and Changes in the Activity of Peptidylarginine Deiminase 3 Induced by High Ca(2+) Concentrations
title_fullStr Autocitrullination and Changes in the Activity of Peptidylarginine Deiminase 3 Induced by High Ca(2+) Concentrations
title_full_unstemmed Autocitrullination and Changes in the Activity of Peptidylarginine Deiminase 3 Induced by High Ca(2+) Concentrations
title_short Autocitrullination and Changes in the Activity of Peptidylarginine Deiminase 3 Induced by High Ca(2+) Concentrations
title_sort autocitrullination and changes in the activity of peptidylarginine deiminase 3 induced by high ca(2+) concentrations
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9386831/
https://www.ncbi.nlm.nih.gov/pubmed/35990454
http://dx.doi.org/10.1021/acsomega.2c02972
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