Zinc transporter 8 (ZnT8) autoantibody epitope specificity and affinity examined with recombinant ZnT8 variant proteins in specific ZnT8R and ZnT8W autoantibody-positive type 1 diabetes patients.
Clin Exp Immunol
; 179(2): 220-9, 2015 Feb.
Article
in En
| MEDLINE
| ID: mdl-25178386
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 type 1 diabetes (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 (i) generate recombinant ZnT8R- and ZnT8W-aa275-369 proteins; (ii) test the ZnT8R- and ZnT8W-aa275-369 proteins in reciprocal competitive radiobinding assays (RBA) against ZnT8R- and ZnT8W-aa268-369 labelled with (35) S-methionine; and (iii) determine the specificity and affinity of sera specific for either ZnT8 arginine (R) or ZnT8 tryptophan (W) autoantibodies in newly diagnosed T1D patients. The results demonstrate, first, that it was possible to produce recombinant human MBP-ZnT8-aa275-369 protein purified to homogeneity for RBA reciprocal competition experiments. Secondly, high-titre ZnT8WA sera diluted to half maximal binding showed significant specificity for respective variants of either ZnT8R or ZnT8W. Thirdly, 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.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Autoantibodies
/
Cation Transport Proteins
/
Diabetes Mellitus, Type 1
/
Antibody Affinity
/
Antibody Specificity
/
Epitopes
Type of study:
Clinical_trials
/
Prognostic_studies
Limits:
Adolescent
/
Adult
/
Child
/
Child, preschool
/
Female
/
Humans
/
Infant
/
Male
Language:
En
Journal:
Clin Exp Immunol
Year:
2015
Document type:
Article
Affiliation country:
Sweden