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ZnT8 autoantibody epitope specificity and affinity examined with recombinant ZnT8 variant proteins in specific ZnT8R and ZnT8W autoantibody positive type 1 diabetes patients.

Author:
  • Hanna Skärstrand
  • Ewa Krupinska
  • Tatu Haataja
  • Fariba Vaziri Sani
  • Jens Lagerstedt
  • Åke Lernmark
Publishing year: 2015
Language: English
Pages: 220-229
Publication/Series: Clinical and Experimental Immunology
Volume: 179
Issue: 2
Document type: Journal article
Publisher: British Society for Immunology

Abstract english

Variant specific zinc transporter 8 autoantibodies (ZnT8A) against either arginine (R) or tryptophan (W) at amino acid (aa) position 325 of the zinc transporter 8 (ZnT8) has been identified in T1D patients. Reciprocal cross-over tests revealed differences in half-maximal binding to indicate variable affinity of patient ZnT8 autoantibodies. Insufficient recombinant ZnT8 variant proteins have precluded detailed analyses of ZnT8 autoantibody affinity. The aims in the present study were to 1) generate recombinant ZnT8R- and ZnT8W-aa275-369 proteins 2) test the ZnT8R- and ZnT8W-aa275-369 proteins in reciprocal competitive radiobinding assays (RBA) against ZnT8R- and ZnT8W-aa268-369 labeled with (35) S-methionine, and 3) determine the specificity and affinity of sera specific for either ZnT8 Arginine (R) or ZnT8 Tryptophan (W) autoantibodies in newly diagnosed type 1 diabetes (T1D) patients. The results demonstrate first that it was possible to produce recombinant human MBP-ZnT8aa275-369 protein purified to homogeneity for RBA reciprocal competition experiments. Second, high titer ZnT8WA sera diluted to half maximal binding showed significant specificity for respective variants of either ZnT8R or ZnT8W. Third, ZnT8WA positive sera showed high affinity for ZnT8W compared to ZnT8RA for ZnT8R. These data demonstrate that T1D patients may have single amino acid specific autoantibodies directed against either ZnT8R or ZnT8W and that the autoantibody affinity to the respective variant may be different. Further studies are needed to assess the mechanisms by which variant specific ZnT8A of variable affinity develop and their possible role in the pathogenic process leading to the clinical onset of T1D.

Keywords

  • Immunology in the medical area

Other

Published
  • Diabetes and Celiac Unit
  • Medical Protein Science
  • Medical Structural Biology
  • ISSN: 0009-9104
E-mail: ake [dot] lernmark [at] med [dot] lu [dot] se

Principal investigator

Diabetes and Celiac Unit

+46 40 39 19 01

+46 70 616 47 79

60:11:015

Jan Waldenströms gata 35, Malmö

33

Lund University Diabetes Centre, CRC, SUS Malmö, Jan Waldenströms gata 35, House 91:12. SE-214 28 Malmö. Telephone: +46 40 39 10 00