ZnT8 autoantibody epitope specificity and affinity examined with recombinant ZnT8 variant proteins in specific ZnT8R and ZnT8W autoantibody positive type 1 diabetes patients.
Skärstrand, Hanna LU ; Krupinska, Ewa LU ; Haataja, Tatu LU ; Vaziri Sani, Fariba LU ; Lagerstedt, Jens LU and Lernmark, Åke LU
(2015)
In Clinical and Experimental Immunology
179(2).
p.220-229
- Abstract
- 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... (More)
- 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. (Less)
Please use this url to cite or link to this publication:
https://lup.lub.lu.se/record/4692625
- author
- Skärstrand, Hanna
LU
; Krupinska, Ewa
LU
; Haataja, Tatu
LU
; Vaziri Sani, Fariba
LU
; Lagerstedt, Jens
LU
and Lernmark, Åke
LU
- organization
- publishing date
- 2015
- type
- Contribution to journal
- publication status
- published
- subject
- in
- Clinical and Experimental Immunology
- volume
- 179
- issue
- 2
- pages
- 220 - 229
- publisher
- British Society for Immunology
- external identifiers
-
- pmid:25178386
- wos:000347233800008
- scopus:84920275601
- pmid:25178386
- ISSN
- 0009-9104
- DOI
- 10.1111/cei.12448
- language
- English
- LU publication?
- yes
- id
- e4b14cf1-29c1-4c57-ae87-78dc5163d2c1 (old id 4692625)
- alternative location
- http://www.ncbi.nlm.nih.gov/pubmed/25178386?dopt=Abstract
- date added to LUP
- 2016-04-01 10:23:20
- date last changed
- 2022-04-27 21:35:14
@article{e4b14cf1-29c1-4c57-ae87-78dc5163d2c1,
abstract = {{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.}},
author = {{Skärstrand, Hanna and Krupinska, Ewa and Haataja, Tatu and Vaziri Sani, Fariba and Lagerstedt, Jens and Lernmark, Åke}},
issn = {{0009-9104}},
language = {{eng}},
number = {{2}},
pages = {{220--229}},
publisher = {{British Society for Immunology}},
series = {{Clinical and Experimental Immunology}},
title = {{ZnT8 autoantibody epitope specificity and affinity examined with recombinant ZnT8 variant proteins in specific ZnT8R and ZnT8W autoantibody positive type 1 diabetes patients.}},
url = {{http://dx.doi.org/10.1111/cei.12448}},
doi = {{10.1111/cei.12448}},
volume = {{179}},
year = {{2015}},
}