DataSheet_1_The first crystal structure of CD8αα from a cartilaginous fish.docx (5.53 MB)
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DataSheet_1_The first crystal structure of CD8αα from a cartilaginous fish.docx

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posted on 2023-04-14, 04:08 authored by Zhao Jia, Jianhua Feng, Helen Dooley, Jun Zou, Junya Wang

Cartilaginous fishes are the most evolutionary-distant vertebrates from mammals and possess an immunoglobulin (Ig)- and T cell-mediated adaptive immunity. CD8 is the hallmark receptor of cytotoxic T cells and is required for the formation of T cell receptor-major histocompatibility complex (TCR-MHC) class I complexes.


RACE PCR was used to obtain gene sequences. Direct dilution was applied for the refolding of denatured recombinant CD8 protein. Hanging-drop vapor diffusion method was performed for protein crystallization.


In this study, CD8α and CD8β orthologues (termed ScCD8α and ScCD8β) were identified in small-spotted catshark (Scyliorhinus canicula). Both ScCD8α and ScCD8β possess an extracellular immunoglobulin superfamily (IgSF) V domain as in previously identified CD8 proteins. The genes encoding CD8α and CD8β are tandemly linked in the genomes of all jawed vertebrates studied, suggesting that they were duplicated from a common ancestral gene before the divergence of cartilaginous fishes and other vertebrates. We determined the crystal structure of the ScCD8α ectodomain homodimer at a resolution of 1.35 Å and show that it exhibits the typical topological structure of CD8α from endotherms. As in mammals, the homodimer formation of ScCD8αα relies upon interactions within a hydrophobic core although this differs in position and amino acid composition. Importantly, ScCD8αα shares the canonical cavity required for interaction with peptide-loaded MHC I in mammals. Furthermore, it was found that ScCD8α can co-immunoprecipitate with ScCD8β, indicating that it can form both homodimeric and heterodimeric complexes.


Our results expand the current knowledge of vertebrate CD8 dimerization and the interaction between CD8α with p/MHC I from an evolutionary perspective.