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Topological assignment of the N-terminal extension of plasma gelsolin to the gelsolin surface.

ABSTRACT: The actin-binding protein gelsolin is highly conserved in vertebrates and exists in two isoforms, a cytoplasmic and an extracellular variant, generated by alternative splicing. In mammals, these isoforms differ only by an N-terminal extension in plasma gelsolin, a short sequence of up to 25 amino acids. Cells and tissues may contain both variants, as plasma gelsolin is secreted by many cell types. The tertiary structure of equine plasma gelsolin has been elucidated, but without any information on the N-terminal extension. In this paper, we present topographical data on the N-terminal extension, derived using a biochemical and immunological approach. For this purpose, a monoclonal antibody was generated that exclusively recognizes cytoplasmic gelsolin but not the extracellular variant and thus allows isoform-specific immunodetection and quantification of cytoplasmic gelsolin in the presence of plasma gelsolin. Using limited proteolysis and pepscan analysis, we mapped the binding epitope and localized it within two regions in segment 1 of the cytoplasmic gelsolin sequence: Tyr34-Ile45 and Leu64-Ile78. In the tertiary structure of the cytoplasmic variant, these sequences are mutually adjacent and located in the proximity of the N-terminus. We therefore conclude that the binding site of the antibody is covered by the N-terminal extension in plasma gelsolin and thus sterically hinders antibody binding. Our results allow for a topological model of the N-terminal extension on the surface of the gelsolin molecule, which was unknown previously.

SUBMITTER: Fock U

PROVIDER: S-EPMC1134740 | biostudies-literature | 2005 Feb

REPOSITORIES: biostudies-literature

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Topological assignment of the N-terminal extension of plasma gelsolin to the gelsolin surface.

Fock Ulrike U Jockusch Brigitte M BM Schubert Wolf-Dieter WD Hinssen Horst H

The Biochemical journal 20050201 Pt 3

The actin-binding protein gelsolin is highly conserved in vertebrates and exists in two isoforms, a cytoplasmic and an extracellular variant, generated by alternative splicing. In mammals, these isoforms differ only by an N-terminal extension in plasma gelsolin, a short sequence of up to 25 amino acids. Cells and tissues may contain both variants, as plasma gelsolin is secreted by many cell types. The tertiary structure of equine plasma gelsolin has been elucidated, but without any information o ...[more]

PMID: 15377282

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Project description:BackgroundExcessive inflammation contributes to the morbidity and mortality of severe coronavirus disease 2019 (COVID-19) pneumonia. Recombinant human plasma gelsolin (rhu-pGSN) improves disease outcomes in diverse experimental models of infectious and noninfectious inflammation.MethodsIn a blinded, randomized study, 61 subjects with documented COVID-19 pneumonia having a World Health Organization (WHO) Severity Score of 4 to 6 and evidence of a hyperinflammatory state were treated with standard care and either adjunctive rhu-pGSN 12 mg/kg or an equal volume of saline placebo given intravenously at entry, 12 hours, and 36 hours. The prespecified coprimary outcomes were survival without major respiratory, hemodynamic, or renal support on Day 14 and the incidence of serious adverse events (SAEs) during the 90-day study period.ResultsAll subjects receiving ≥1 dose of study drug were analyzed. Fifty-four of 61 subjects (88.5%) were WHO severity level 4 at entry. The proportions of subjects alive without support on Day 14 were 25 of 30 rhu-pGSN recipients (83.3%) and 27 of 31 placebo recipients (87.1%). Over the duration of the study, WHO Severity Scores improved similarly in both treatment groups. No statistically significant differences were observed between treatment groups at any time point examined. Two subjects died in each group. Numerically fewer subjects in the rhu-pGSN group had SAEs (5 subjects; 16.7%) or ≥ Grade 3 adverse events (5 subjects; 16.7%) than in the placebo group (8 subjects [25.8%] and 9 subjects [29.0%], respectively), mostly involving the lungs. Three rhu-pGSN recipients (10.0%) were intubated compared to 6 placebo recipients (19.4%).ConclusionsOverall, subjects in this study did well irrespective of treatment arm. When added to dexamethasone and remdesivir, no definitive benefit was demonstrated for rhu-pGSN relative to placebo. Safety signals were not identified after the administration of 3 doses of 12 mg/kg rhu-pGSN over 36 hours. The frequencies of SAEs and intubation were numerically fewer in the rhu-pGSN group compared with placebo.

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Topological assignment of the N-terminal extension of plasma gelsolin to the gelsolin surface.

Publications

Topological assignment of the N-terminal extension of plasma gelsolin to the gelsolin surface.

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