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Structural and functional insights into Escherichia coli ?2-macroglobulin endopeptidase snap-trap inhibition.

ABSTRACT: The survival of commensal bacteria requires them to evade host peptidases. Gram-negative bacteria from the human gut microbiome encode a relative of the human endopeptidase inhibitor, ?2-macroglobulin (?2M). Escherichia coli ?2M (ECAM) is a ? 180-kDa multidomain membrane-anchored pan-peptidase inhibitor, which is cleaved by host endopeptidases in an accessible bait region. Structural studies by electron microscopy and crystallography reveal that this cleavage causes major structural rearrangement of more than half the 13-domain structure from a native to a compact induced form. It also exposes a reactive thioester bond, which covalently traps the peptidase. Subsequently, peptidase-laden ECAM is shed from the membrane and may dimerize. Trapped peptidases are still active except against very large substrates, so inhibition potentially prevents damage of large cell envelope components, but not host digestion. Mechanistically, these results document a novel monomeric "snap trap."

SUBMITTER: Garcia-Ferrer I

PROVIDER: S-EPMC4500212 | biostudies-literature | 2015 Jul

REPOSITORIES: biostudies-literature

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Structural and functional insights into Escherichia coli α2-macroglobulin endopeptidase snap-trap inhibition.

Garcia-Ferrer Irene I Arêde Pedro P Gómez-Blanco Josué J Luque Daniel D Duquerroy Stephane S Castón José R JR Goulas Theodoros T Gomis-Rüth F Xavier FX

Proceedings of the National Academy of Sciences of the United States of America 20150622 27

The survival of commensal bacteria requires them to evade host peptidases. Gram-negative bacteria from the human gut microbiome encode a relative of the human endopeptidase inhibitor, α2-macroglobulin (α2M). Escherichia coli α2M (ECAM) is a ∼ 180-kDa multidomain membrane-anchored pan-peptidase inhibitor, which is cleaved by host endopeptidases in an accessible bait region. Structural studies by electron microscopy and crystallography reveal that this cleavage causes major structural rearrangemen ...[more]

PMID: 26100869

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Project description:Advanced osteoradionecrosis (ORN) is one of the most serious complications in patients with head and neck cancer, resulting in poor prognosis. Numerous studies have therefore focused on the pathogenesis and interventions of ORN early stage. The present study aimed to investigate whether α2‑macroglobulin (α2M) could prevent early‑stage jaw osteoradionecrosis caused by radiotherapy (RT). Following local injection of α2M, a single dose of 30 Gy was delivered to rats for pathological exploration. For 28 days, the irradiated mandible and soft tissues were examined for potential changes. Furthermore, primary human bone marrow mesenchymal stem cells pretreated with α2M followed by 8 Gy irradiation (IR) were also used. Tartrate‑resistant acid phosphatase assay, terminal uridine deoxynucleotidyl nick end labeling assay and immunohistochemical staining were performed on irradiated mandibular bone, tongue or buccal mucosa tissues from rats. Cell proliferation was assessed by evaluating the cell morphology by microscopy and by using the cell counting kit‑8. Fluorescence staining, flow cytometry and western blotting were conducted to detect the reactive oxygen species level, cell apoptosis and protein expression of superoxide dismutase 2 (SOD2), heme oxygenase‑1 (HO‑1) and phosphorylated Akt following irradiation. The results demonstrated that α2M attenuated physical inflammation, osteoclasts number and fat vacuole accumulation in mandibular bone marrow and bone marrow cell apoptosis following IR in vivo. Furthermore, α2M pretreatment suppressed the expression of 8‑hydroxy‑2'‑deoxyguanosine in mandibular bone and tongue paraffin embedded sections, which is a marker of oxidative damage, and increased SOD2 expression in mucosa and tongue paraffin embedded sections. The present study demonstrated the efficient regulation of antioxidative enzymes, including SOD2 and heme oxygenase‑1, and reduction in oxidative damage by α2M. In addition, in vitro results confirmed that α2M may protect cells from apoptosis and suppress reactive oxygen species accumulation. Overall, the present study demonstrated that α2M treatment may exert some radioprotective effects in early‑stage ORN via antioxidant mechanisms, and may therefore be considered as a potential alternative molecule in clinical prophylactic treatments.

Dataset Information

Structural and functional insights into Escherichia coli ?2-macroglobulin endopeptidase snap-trap inhibition.

Publications

Structural and functional insights into Escherichia coli α2-macroglobulin endopeptidase snap-trap inhibition.

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