Identification and quantification of sites of nitration and oxidation in the key matrix protein laminin and the structural consequences of these modifications

Lasse G. Lorentzen; Christine Y. Chuang; Adelina Rogowska-Wrzesinska; Michael J. Davies

doi:10.1016/j.redox.2019.101226

Identification and quantification of sites of nitration and oxidation in the key matrix protein laminin and the structural consequences of these modifications

Lasse G. Lorentzen, Christine Y. Chuang, Adelina Rogowska-Wrzesinska, Michael J. Davies

Inflammation, Metabolism and Oxidation

4 Citationer (Scopus)

Abstract

Laminin is a major protein of the basement membrane (BM), a specialized extracellular matrix (ECM) of the artery wall. The potent oxidizing and nitrating agent peroxynitrous acid (ONOOH) is formed at sites of inflammation, and data implicate ONOOH in ECM damage and cardiovascular disease. Co-localization of 3-nitrotyrosine, a product of ONOOH-mediated tyrosine (Tyr) modification, and laminin has been reported in human atherosclerotic lesions. The sites and consequences of 3-nitrotyrosine (and related nitrated tryptophan) formation on laminin, ifs self-assembly and cell interactions are poorly understood. In this study murine laminin-111 was exposed to ONOOH (1-500-fold molar excess). Nitration sites were mapped and quantified using LC-MS/MS. Mono-nitration was detected at 148 sites (126 Tyr, 22 Trp), and di-nitration at 14 sites. Label-free quantification showed enhanced nitration with increasing oxidant doses. Tyr nitration was similar to 10-fold greater than at Trp. CO2 modulated damage in a site-specific manner, with most sites less extensively nitrated. 119 mononitration sites were identified with CO2 present, and no unique sites were detected. 23 di-nitration sites were detected, with 15 unique to the presence of CO2. Extensive modification was detected at sites involved in cell adhesion, protein-protein interactions and self-polymerization. Tyr-145 on the gamma 1 chain was extensively nitrated, and endothelial cells exhibited decreased adhesion to a nitrated peptide modelling this site. Modification of residues involved in self-polymerization interfered with the formation of ordered polymers as detected by scanning electron microscopy. These laminin modifications may contribute to endothelial cell dysfunction and modulate ECM structure and assembly, and thereby contribute to atherogenesis.

Originalsprog	Engelsk
Artikelnummer	UNSP 101226
Tidsskrift	Redox Biology
Vol/bind	24
ISSN	2213-2317
DOI	https://doi.org/10.1016/j.redox.2019.101226
Status	Udgivet - jun. 2019

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10.1016/j.redox.2019.101226Licens: CC BY-NC-ND

http://europepmc.org/articles/pmc6543125?pdf=renderLicens: CC BY-NC-ND

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Identification and quantification of sites of nitration and oxidation in the key matrix protein laminin and the structural consequences of these modifications. / Lorentzen, Lasse G.; Chuang, Christine Y.; Rogowska-Wrzesinska, Adelina et al.

I: Redox Biology, Bind 24, UNSP 101226, 06.2019.

Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › peer review

@article{4b50eeda62314e17a913bc4cb2919fd2,

title = "Identification and quantification of sites of nitration and oxidation in the key matrix protein laminin and the structural consequences of these modifications",

abstract = "Laminin is a major protein of the basement membrane (BM), a specialized extracellular matrix (ECM) of the artery wall. The potent oxidizing and nitrating agent peroxynitrous acid (ONOOH) is formed at sites of inflammation, and data implicate ONOOH in ECM damage and cardiovascular disease. Co-localization of 3-nitrotyrosine, a product of ONOOH-mediated tyrosine (Tyr) modification, and laminin has been reported in human atherosclerotic lesions. The sites and consequences of 3-nitrotyrosine (and related nitrated tryptophan) formation on laminin, ifs self-assembly and cell interactions are poorly understood. In this study murine laminin-111 was exposed to ONOOH (1-500-fold molar excess). Nitration sites were mapped and quantified using LC-MS/MS. Mono-nitration was detected at 148 sites (126 Tyr, 22 Trp), and di-nitration at 14 sites. Label-free quantification showed enhanced nitration with increasing oxidant doses. Tyr nitration was similar to 10-fold greater than at Trp. CO2 modulated damage in a site-specific manner, with most sites less extensively nitrated. 119 mononitration sites were identified with CO2 present, and no unique sites were detected. 23 di-nitration sites were detected, with 15 unique to the presence of CO2. Extensive modification was detected at sites involved in cell adhesion, protein-protein interactions and self-polymerization. Tyr-145 on the gamma 1 chain was extensively nitrated, and endothelial cells exhibited decreased adhesion to a nitrated peptide modelling this site. Modification of residues involved in self-polymerization interfered with the formation of ordered polymers as detected by scanning electron microscopy. These laminin modifications may contribute to endothelial cell dysfunction and modulate ECM structure and assembly, and thereby contribute to atherogenesis.",

keywords = "Extracellular matrix, Peroxynitrous acid, Peroxynitrite, Laminin, Protein oxidation, 3-Nitrotyrosine",

author = "Lorentzen, {Lasse G.} and Chuang, {Christine Y.} and Adelina Rogowska-Wrzesinska and Davies, {Michael J.}",

year = "2019",

month = jun,

doi = "10.1016/j.redox.2019.101226",

language = "English",

volume = "24",

journal = "Redox Biology",

issn = "2213-2317",

publisher = "Elsevier",

}

TY - JOUR

T1 - Identification and quantification of sites of nitration and oxidation in the key matrix protein laminin and the structural consequences of these modifications

AU - Lorentzen, Lasse G.

AU - Chuang, Christine Y.

AU - Rogowska-Wrzesinska, Adelina

AU - Davies, Michael J.

PY - 2019/6

Y1 - 2019/6

N2 - Laminin is a major protein of the basement membrane (BM), a specialized extracellular matrix (ECM) of the artery wall. The potent oxidizing and nitrating agent peroxynitrous acid (ONOOH) is formed at sites of inflammation, and data implicate ONOOH in ECM damage and cardiovascular disease. Co-localization of 3-nitrotyrosine, a product of ONOOH-mediated tyrosine (Tyr) modification, and laminin has been reported in human atherosclerotic lesions. The sites and consequences of 3-nitrotyrosine (and related nitrated tryptophan) formation on laminin, ifs self-assembly and cell interactions are poorly understood. In this study murine laminin-111 was exposed to ONOOH (1-500-fold molar excess). Nitration sites were mapped and quantified using LC-MS/MS. Mono-nitration was detected at 148 sites (126 Tyr, 22 Trp), and di-nitration at 14 sites. Label-free quantification showed enhanced nitration with increasing oxidant doses. Tyr nitration was similar to 10-fold greater than at Trp. CO2 modulated damage in a site-specific manner, with most sites less extensively nitrated. 119 mononitration sites were identified with CO2 present, and no unique sites were detected. 23 di-nitration sites were detected, with 15 unique to the presence of CO2. Extensive modification was detected at sites involved in cell adhesion, protein-protein interactions and self-polymerization. Tyr-145 on the gamma 1 chain was extensively nitrated, and endothelial cells exhibited decreased adhesion to a nitrated peptide modelling this site. Modification of residues involved in self-polymerization interfered with the formation of ordered polymers as detected by scanning electron microscopy. These laminin modifications may contribute to endothelial cell dysfunction and modulate ECM structure and assembly, and thereby contribute to atherogenesis.

AB - Laminin is a major protein of the basement membrane (BM), a specialized extracellular matrix (ECM) of the artery wall. The potent oxidizing and nitrating agent peroxynitrous acid (ONOOH) is formed at sites of inflammation, and data implicate ONOOH in ECM damage and cardiovascular disease. Co-localization of 3-nitrotyrosine, a product of ONOOH-mediated tyrosine (Tyr) modification, and laminin has been reported in human atherosclerotic lesions. The sites and consequences of 3-nitrotyrosine (and related nitrated tryptophan) formation on laminin, ifs self-assembly and cell interactions are poorly understood. In this study murine laminin-111 was exposed to ONOOH (1-500-fold molar excess). Nitration sites were mapped and quantified using LC-MS/MS. Mono-nitration was detected at 148 sites (126 Tyr, 22 Trp), and di-nitration at 14 sites. Label-free quantification showed enhanced nitration with increasing oxidant doses. Tyr nitration was similar to 10-fold greater than at Trp. CO2 modulated damage in a site-specific manner, with most sites less extensively nitrated. 119 mononitration sites were identified with CO2 present, and no unique sites were detected. 23 di-nitration sites were detected, with 15 unique to the presence of CO2. Extensive modification was detected at sites involved in cell adhesion, protein-protein interactions and self-polymerization. Tyr-145 on the gamma 1 chain was extensively nitrated, and endothelial cells exhibited decreased adhesion to a nitrated peptide modelling this site. Modification of residues involved in self-polymerization interfered with the formation of ordered polymers as detected by scanning electron microscopy. These laminin modifications may contribute to endothelial cell dysfunction and modulate ECM structure and assembly, and thereby contribute to atherogenesis.

KW - Extracellular matrix

KW - Peroxynitrous acid

KW - Peroxynitrite

KW - Laminin

KW - Protein oxidation

KW - 3-Nitrotyrosine

U2 - 10.1016/j.redox.2019.101226

DO - 10.1016/j.redox.2019.101226

M3 - Journal article

C2 - 31154162

SN - 2213-2317

VL - 24

JO - Redox Biology

JF - Redox Biology

M1 - UNSP 101226

ER -

Identification and quantification of sites of nitration and oxidation in the key matrix protein laminin and the structural consequences of these modifications

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