Globular-shaped variable lymphocyte receptors B antibody multimerized by a hydrophobic clustering in hagfish

Jaesung Kim,Kyun Oh Lee,Gwang Joong Kim,Kim D Thompson,Jong Yong Kim,Tae Sung Jung,Si Won Kim,Young Rim Kim,Se Pyeong Im,Jae Wook Jung,Jassy Mary S Lazarte,Alexandra Adams,Sangmin Lee,Hyun Suk Jung,Jung Seok Lee

doi:10.1038/s41598-018-29197-w

Abstract

In hagfish and lampreys, two representative jawless vertebrates, the humoral immunity is directly mediated by variable lymphocyte receptors B (VLRBs). Both monomeric VLRBs are structurally and functionally similar, but their C-terminal tails differ: lamprey VLRB has a Cys-rich tail that forms disulfide-linked pentamers of dimers, contributing to its multivalency, whereas hagfish VLRB has a superhydrophobic tail of unknown structure. Here, we reveal that VLRBs obtained from hagfish plasma have a globular-shaped multimerized form (approximately 0.6 to 1.7 MDa) that is generated by hydrophobic clustering instead of covalent linkage. Electron microscopy (EM) and single-particle analysis showed that the multimerized VLRBs form globular-shaped clusters with an average diameter of 28.7 ± 2.2 nm. The presence of VLRBs in the complex was confirmed by immune-EM analysis using an anti-VLRB antibody. Furthermore, the hydrophobic hagfish C-terminus (HC) was capable of triggering multimerization and directing the cellular surface localization via a glycophosphatidylinositol linkage. Our results strongly suggest that the hagfish VLRB forms a previously unknown globular-shaped antibody. This novel identification of a structurally unusual VLRB complex may suggest that the adaptive immune system of hagfish differs from that of lamprey.

Highlights

In hagfish and lampreys, two representative jawless vertebrates, the humoral immunity is directly mediated by variable lymphocyte receptors B (VLRBs)
Albeit weaker, dose-dependent disruptions of the VLRB complex were observed following treatment with non-ionic detergents, such as NP-40 or Triton X-100 (Fig. 1C). As these detergents are commonly used to denature proteins by disrupting non-covalent interaction and permeating into the hydrophobic core of native proteins[16,17], these results suggest that circulating hagfish VLRB may naturally exist as multimeric complexes that lack covalent linkages, such as the disulfide bonds formed by the lamprey C-terminus (LC) in lamprey
Our results demonstrate that individual VLRB monomers could be tightly packed together by self-assembly via hydrophobic clustering of the hagfish C-terminus (HC)

Summary

Introduction

Two representative jawless vertebrates, the humoral immunity is directly mediated by variable lymphocyte receptors B (VLRBs). Since the middle of the 20th century, results published by various researchers have shown that Ag-specific agglutinins and immunological memory can be produced by the AISs of lampreys and hagfish[9,10] Such studies showed that fish immunized with various types of Ags could generate Ag-specific agglutinin molecules with molecular weights similar to that of an immunoglobulin (Ig) M antibody. This Ag-specific agglutination was explained by the discovery that lamprey VLRB is capable of multimerization[11]. We for the first time describe a unique structure of multimerized VLRB antibodies in hagfish plasma and demonstrate that the multimerization of hagfish VLRB could be mediated by hydrophobic clustering directed by its C-terminal region

Methods

Results

Conclusion