Functional importance of covalent homodimer of reelin protein linked via its central region

Norihisa Yasui, Yu Kitago, Ayako Beppu, Takao Kohno, Shunsuke Morishita, Hiroki Gomi, Masamichi Nagae, Mitsuharu Hattori, Junichi Takagi

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16 Citations (Scopus)

Abstract

Reelin is a 3461-residue secreted glycoprotein that plays a critical role in brain development through its action on target neurons. Although it is known that functional reelin protein exists as multimer formed by interchain disulfide bond(s) as well as through non-covalent interactions, the chemical nature of the multimer assembly has been elusive. In the present study, we identified, among 122 cysteines present in full-length reelin, the single critical cysteine residue (Cys 2101) responsible for the covalent multimerization. C2101A mutant reelin failed to assemble into disulfide-bonded multimers, whereas it still exhibited non-covalently associated high molecular weight oligomeric states in solution. Detailed analysis of tryptic fragments produced from the purified reelin proteins revealed that the minimum unit of the multimer is a homodimeric reelin linked via Cys 2101 present in the central region and that this cysteine does not connect to the N-terminal region of reelin, which had been postulated as the primary oligomerization domain. A surface plasmon resonance binding assay confirmed that C2101A mutant reelin retained binding capability toward two neuronal receptors apolipoprotein E receptor 2 and very low density lipoprotein receptor. However, it failed to show signaling activity in the assay using the cultured neurons. These results indicate that an intact higher order architecture of reelin multimer maintained by both Cys 2101-mediated homodimerization and other non-covalent association present elsewhere in the reelin primary structure are essential for exerting its full biological activity.

Original languageEnglish
Pages (from-to)35247-35256
Number of pages10
JournalJournal of Biological Chemistry
Volume286
Issue number40
DOIs
Publication statusPublished - Oct 7 2011
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Yasui, N., Kitago, Y., Beppu, A., Kohno, T., Morishita, S., Gomi, H., Nagae, M., Hattori, M., & Takagi, J. (2011). Functional importance of covalent homodimer of reelin protein linked via its central region. Journal of Biological Chemistry, 286(40), 35247-35256. https://doi.org/10.1074/jbc.M111.242719