Conformation-dependent GAD65 autoantibodies in diabetes

Luo, Dong; Gilliam, Lisa K.; Greenbaum, Carla J.; Bekris, Lynn M.; Hampe, Christiane S.; Daniels, Terri; Richter, Wiltrud; Marcovina, Santica M.; Rolandsson, Olov; Landin‐Olsson, Mona; Kockum, Ingrid; Lernmark, Åke

doi:10.1007/s00125-004-1495-3

Cited by 7 publications

(4 citation statements)

References 49 publications

(70 reference statements)

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“…Thus, it remains to be determined whether these peptides represent natural epitopes related to SPS or alternatively T1D. A few studies describe that the major epitopes of GAD67 appears to be conformational, similar to the major epitopes of GAD65 . Thus, identification of these N ‐terminal continuous epitopes of GAD67 may be valuable in relation to development of potential therapeutic reagents.…”

Section: Discussionmentioning

confidence: 99%

“…Analysis of GAD autoantibodies in SPS and T1D has shown similarities and differences in relation to antigen recognition . In both diseases, GAD antibody reactivity is predominantly directed to conformational epitope regions in the middle and the C ‐terminal domains of GAD65 (amino acids 242–439 and 450–585) . Additionally in SPS, there is reactivity to conformational epitopes on GAD67 , and short linear epitopes in the C ‐terminal region and in the N ‐terminus of GAD65 .…”

Section: Introductionmentioning

confidence: 99%

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Characterization of continuous B‐cell epitopes in the N‐terminus of glutamate decarboxylase67 using monoclonal antibodies

Agca

Trier

2014

Journal of Peptide Science

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Glutamate decarboxylase (GAD) is an autoantigen associated with the autoimmune disorders Type-1 diabetes (T1D) and stiff-person syndrome (SPS). The protein, being an essential enzyme involved in the production of the inhibitory neurotransmitter γ-aminobutyric acid, exists in two isoforms, GAD67 and GAD65. Both isoforms may be targeted by autoantibodies in SPS and T1D patients, although SPS primarily is associated with the presence of GAD67 autoantibodies, whereas T1D mainly is associated with the presence of GAD65 autoantibodies. In this study, we describe antibody reactivity to overlapping GAD67 peptides covering the complete protein sequence by modified peptide enzyme-linked immunosorbent assay in order to identify potential GAD67 epitopes using two monoclonal antibodies (mAbs). Both GAD67 mAbs showed reactivity to linear epitopes located at the N-terminal end of GAD67. The epitopes of GAD mAb 1 and 2 were identified as the amino acid sequences NAGADPNTTN and TETDFSNLF, respectively, corresponding to amino acids 14-23 and 91-99. Fine mapping of the epitopes revealed that antibody reactivity was related to amino acid side-chain functionality, rather than amino acid side-chain specificity. Additionally, results suggested that non-contact amino acids in the epitope structure were essential for antibody reactivity. The exact role of these amino acids remains to be determined, but they are thought to be involved in backbone hydrogen bonds or stabilization of the epitope structure. As only limited knowledge is available in relation to antigenic regions of GAD67, this study contributes to characterization of GAD67 epitopes and may be a first step in the development of peptide-based therapeutics against SPS.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Characterization of continuous B‐cell epitopes in the N‐terminus of glutamate decarboxylase67 using monoclonal antibodies

Agca

Trier

2014

Journal of Peptide Science

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“…The two isoforms are divided into three functional domains, an amino-terminal domain, a middle domain where the catalytic site resides (PLP-binding domain), and a carboxy-terminal domain. Roughly, antibodies to GAD67 are primarily associated with stiff-person syndrome (SPS) and autoimmune polyendocrine syndrome type 1, whereas GAD65 is a dominant autoantigen in type 1 diabetes (T1D) [2,24,25,26,27,28,29].…”

Section: Introductionmentioning

confidence: 99%

Fine Mapping of Glutamate Decarboxylase 65 Epitopes Reveals Dependency on Hydrophobic Amino Acids for Specific Interactions

Valdarnini

Holm

Hansen

et al. 2019

IJMS

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Characterization of multiple antibody epitopes has revealed the necessity of specific groups of amino acid residues for reactivity. This applies to the majority of antibody–antigen interactions, where especially charged and hydrophilic amino acids have been reported to be essential for antibody reactivity. This study describes thorough characterization of glutamic acid decarboxylase (GAD) 65 antigenic epitopes, an immunodominant autoantigen in type 1 diabetes (T1D). As linear epitopes are sparsely described for GAD65 in T1D, we aimed to identify and thoroughly characterize two GAD65 antibodies using immunoassays. A monoclonal antibody recognized an epitope in the N-terminal domain of GAD65, 8FWSFGSE14, whereas a polyclonal antibody recognized two continuous epitopes in the C-terminal domain, corresponding to amino acids 514RTLED518 and 549PLGDKVNF556. Hydrophobic amino acids were essential for antibody reactivity, which was verified by competitive inhibition assays. Moreover, the epitopes were located in flexible linker regions and turn structures. These findings confirm the versatile nature of antibody–antigen interactions and describe potential continuous epitopes related to T1D, which predominantly have been proposed to be of discontinuous nature.

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“…The two GAD isoforms differ in their enzyme activity and localization. GAD67 is primarily located in the cytoplasm, is constitutively active, and provides for the steady basal production of GABA, whereas GAD65 mainly is present in synaptic vesicles, undergoes auto-inactivation during enzyme activity, and occurs in the cell, providing for a pulse in production under circumstances that demand a rapid surge of GABA synthesis and release [14,18,19]. The two isoforms show high sequence similarity, with the middle and C-terminal domains having 74% identity but differing (25% identity) in the N-terminal domain, mainly in the first 100 amino acids [10,13,14].…”

Section: Introductionmentioning

confidence: 99%

Peptide Antibody Reactivity to Homologous Regions in Glutamate Decarboxylase Isoforms and Coxsackievirus B4 P2C

Trier

Valdarnini

Fanelli

et al. 2022

IJMS

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Two isoforms of the glutamate decarboxylase (GAD) enzyme exist, GAD65 and GAD67, which are associated with type 1 diabetes (T1D) and stiff-person syndrome (SPS), respectively. Interestingly, it has been reported that T1D patients seldom develop SPS, whereas patients with SPS occasionally develop T1D. In addition, coxsackievirus B4 (CVB4) has previously been proposed to be involved in the onset of T1D through molecular mimicry. On this basis, we aimed to examine antibody cross-reactivity between a specific region of GAD65 and GAD67, which has high sequence homology to the nonstructural P2C protein of CVB4 to determine potential correlations at antibody level. Monoclonal peptide antibodies generated in mice specific for a region with high similarity in all three proteins were screened for reactivity along with human sera in immunoassays. In total, six antibodies were generated. Two of the antibodies reacted to both GAD isoforms. However, none of the antibodies were cross-reactive to CVB, suggesting that antibody cross-reactivity between GAD65 and CVB, and GAD67 and CVB may not contribute to the onset of T1D and SPS, respectively.

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Conformation-dependent GAD65 autoantibodies in diabetes

Cited by 7 publications

References 49 publications

Characterization of continuous B‐cell epitopes in the N‐terminus of glutamate decarboxylase67 using monoclonal antibodies

Characterization of continuous B‐cell epitopes in the N‐terminus of glutamate decarboxylase67 using monoclonal antibodies

Fine Mapping of Glutamate Decarboxylase 65 Epitopes Reveals Dependency on Hydrophobic Amino Acids for Specific Interactions

Peptide Antibody Reactivity to Homologous Regions in Glutamate Decarboxylase Isoforms and Coxsackievirus B4 P2C

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