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The Biochemical journal2004; 385(Pt 3); 659-665; doi: 10.1042/BJ20040875

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.
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Publication Date: 2004-09-21 PubMed ID: 15377282PubMed Central: PMC1134740DOI: 10.1042/BJ20040875Google Scholar: Lookup
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  • Journal Article
  • Research Support
  • Non-U.S. Gov't

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

This research focuses on studying more about the actin-binding protein gelsolin, particularly, the vertebrates’ plasma gelsolin by identifying its N-terminal sequence’s positioning. The whole process was found out through a biochemical and immunological approach, and a model of it was created.

Understanding The Research

  • The research starts by talking about gelsolin, a protein that binds with actin. Scattered widely in the animal kingdom, this protein comes in two versions, a cytoplasmic one and an extracellular one.
  • The difference between the two is that the plasma gelsolin – an extracellular variant, comes with an N-terminal extension. This feature makes it secreted by many cell types, thus equally present in cells and tissues with cytoplasmic gelsolin.
  • While the tertiary structure of plasma gelsolin has been exposed, information about the N-terminal extension was missing.

Methodological Approach

  • To comprehend the structure of the N-terminal extension, the study utilizes methods that include biochemical and immunological approaches.
  • A special monoclonal antibody is formulated for this purpose that can only recognize the cytoplasmic gelsolin but not the extracellular one. The antibody thus enables the study to immunodetect and quantify cytoplasmic gelsolin in the company of plasma gelsolin.

Findings

  • Through limited proteolysis and pepscan analysis, researchers were able to map the binding epitope and locate it within two regions in the cytoplasmic gelsolin sequence: Tyr34-Ile45 and Leu64-Ile78.
  • In the cytoplasmic variant’s tertiary structure, these sequences are mutually adjacent and lie near the N-terminus.
  • The inference is that the antibody’s binding site gets covered by the N-terminal extension in plasma gelsolin, thus preventing antibody binding.
  • The research thus manages to ascertain a topological model of the N-terminal extension on the gelsolin molecule’s surface, which wasn’t known earlier.

Cite This Article

APA
Fock U, Jockusch BM, Schubert WD, Hinssen H. (2004). Topological assignment of the N-terminal extension of plasma gelsolin to the gelsolin surface. Biochem J, 385(Pt 3), 659-665. https://doi.org/10.1042/BJ20040875

Publication

The Biochemical journal
ISSN: 1470-8728
NlmUniqueID: 2984726R
Country: England
Language: English
Volume: 385
Issue: Pt 3
Pages: 659-665

Researcher Affiliations

Fock, Ulrike
  • Cell Biology, Zoological Institute, Technical University of Braunschweig, D-38106 Braunschweig, Germany.
Jockusch, Brigitte M
    Schubert, Wolf-Dieter
      Hinssen, Horst

        MeSH Terms

        • Amino Acid Sequence
        • Animals
        • Antibodies, Monoclonal / immunology
        • Antibody Specificity
        • Binding Sites
        • Cytoplasm / metabolism
        • Enzyme-Linked Immunosorbent Assay
        • Epitope Mapping
        • Epitopes / chemistry
        • Epitopes / immunology
        • Epitopes / metabolism
        • Gelsolin / chemistry
        • Gelsolin / genetics
        • Gelsolin / immunology
        • Gelsolin / metabolism
        • Genetic Variation
        • Humans
        • Models, Molecular
        • Molecular Sequence Data
        • Protein Conformation
        • Sensitivity and Specificity
        • Swine

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        Citations

        This article has been cited 1 times.
        1. Piktel E, Levental I, Durnaś B, Janmey PA, Bucki R. Plasma Gelsolin: Indicator of Inflammation and Its Potential as a Diagnostic Tool and Therapeutic Target. Int J Mol Sci 2018 Aug 25;19(9).
          doi: 10.3390/ijms19092516pubmed: 30149613google scholar: lookup
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