Autoreactivity of human VH domains from cDNA libraries: Analysis with a bacterial expression system

Jean Michel Lecerf, Yan Chen, Pascale Richalet-Sécordel, Xiaowei Wang, B. David Stollar

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13 Scopus citations

Abstract

Previous studies showed that VH domains of several anti-DNA Abs can bind DNA in the absence of VL. In the current work, we tested the VH autoreactive potential more generally, examining VH domains that did not come from known autoantibodies. Using a bacterial expression system, we produced 11 fusion proteins, each containing a VH domain and a B domain of staphylococcal protein A. The VH domains were coded in cDNA libraries from circulating B cells of healthy young adult humans. Thus, binding properties of the Ig molecules from which they came were unknown. The B cells had not been stimulated in vitro. Seven cDNA clones combined the frequently expressed V(H)3-23 gene segment with varied D(H) and J(H) segments. The other clones contained unmutated V(H)3-7, V(H)3-9, V(H)3-53, and V(H)4-39 segments. We compared these bacterial expression products with single-chain F(V), VH and VL domains of IgM mAb 18/2, a V(H)3-23-encoded, DNA-binding autoantibody. Submicromolar concentrations of 5 of the 11 VH domains bound to ssDNA. Those and one more also bound to immobilized poly(dT), and two bound to circular plasmid dsDNA. Soluble poly(dT) was the most potent inhibitor in competitive ELISA. Seven of the VII domains also bound to immobilized nuclear ribonucleoprotein, four to histone and none to thyroglobulin. Two interacted with the matrix of a Sephacryl S-100 column. The polyreactive autoantigen- binding properties of these VH domains raise the question of whether these properties may play a role in the formation of the VH repertoire of circulating B cells.

Original languageEnglish
Pages (from-to)1274-1283
Number of pages10
JournalJournal of Immunology
Volume161
Issue number3
StatePublished - Aug 1 1998
Externally publishedYes

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    Lecerf, J. M., Chen, Y., Richalet-Sécordel, P., Wang, X., & David Stollar, B. (1998). Autoreactivity of human VH domains from cDNA libraries: Analysis with a bacterial expression system. Journal of Immunology, 161(3), 1274-1283.