Effect of Rothia mucilaginosa enzymes on gliadin (gluten) structure, deamidation, and immunogenic epitopes relevant to celiac disease.
Effect of Rothia mucilaginosa enzymes on gliadin (gluten) structure, deamidation, and immunogenic epitopes relevant to celiac disease.
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Rothia mucilaginosa 酶对与乳糜泻相关的麦醇溶蛋白（麸质）结构、脱酰胺和免疫原性表位的影响。
DOI：
10.1152/ajpgi.00144.2014
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发表时间：
2014
期刊：
影响因子：
--
通讯作者：
Helmerhorst,EvaJ
中科院分区：
文献类型：
--
作者：
Tian,Na;Wei,Guoxian;Schuppan,Detlef;Helmerhorst,EvaJ
粘液罗氏菌是口腔中的一种天然微生物，它能在抗哺乳动物酶降解的区域切割谷蛋白（麦醇溶蛋白）。本研究的目的是调查在何种程度上R。与粘液素细胞相关的酶消除了麦胶蛋白的免疫原性。通过SDS-PAGE和RP-HPLC监测总麦醇溶蛋白和高免疫原性麦醇溶蛋白33-mer或26-mer肽的降解，并通过液相色谱和电喷雾串联质谱（LC-ESI-MS/MS）对片段进行测序。通过组织转氨酶（TG 2）的肽脱酰胺（使片段更具免疫原性的关键步骤）通过TG 2介导的与单丹酰尸胺（MDC）的交联以及通过LC-ESI-MS的+1-Da质量差来评估。与粘蛋白相关的酶将麦醇溶蛋白、33-mer和26-mer肽切割成更小的片段。TG 2介导的交联与完整的33-mer和26-mer肽水平呈完美的反比关系，主要降解片段的MDC交联速率较慢，为6.18 ± 2.20 Au/min，而完整的33-mer和26-mer分别为97.75 ± 10.72和84.17 ± 3.25 Au/min，这通过在质谱分析中还原的TG 2介导的片段脱酰胺得到证实。麦醇溶蛋白与Rothiacells孵育30 min、2 h和5 h后，R5抗体结合率分别降低了20、82和97%，这与酶处理的33-mer和26-mer肽在R5竞争ELISA中的反应性降低有关。我们广泛的互补方法来验证谷蛋白降解活性qualifiesR。粘液相关酶作为中和T细胞免疫原性的有希望的工具用于治疗乳糜泻。
Rothia mucilaginosa, a natural microbial inhabitant of the oral cavity, cleaves gluten (gliadin) proteins at regions that are resistant to degradation by mammalian enzymes. The aim of this study was to investigate to what extent theR. mucilaginosacell-associated enzymes abolish gliadin immunogenic properties. Degradation of total gliadins and highly immunogenic gliadin 33-mer or 26-mer peptides was monitored by SDS-PAGE and RP-HPLC, and fragments were sequenced by liquid chromatography and electrospray ionization tandem mass spectrometer (LC-ESI-MS/MS). Peptide deamidation by tissue transglutaminase (TG2), a critical step in rendering the fragments more immunogenic, was assessed by TG2-mediated cross-linking to monodansyl cadaverine (MDC), and by a +1-Da mass difference by LC-ESI-MS. Survival of potential immunogenic gliadin epitopes was determined by use of the R5 antibody-based ELISA.R. mucilaginosa-associated enzymes cleaved gliadins, 33-mer and 26-mer peptides into smaller fragments. TG2-mediated cross-linking showed a perfect inverse relationship with intact 33-mer and 26-mer peptide levels, and major degradation fragments showed a slow rate of MDC cross-linking of 6.18 ± 2.20 AU/min compared with 97.75 ± 10.72 and 84.17 ± 3.25 AU/min for the intact 33-mer and 26-mer, respectively, which was confirmed by reduced TG2-mediated deamidation of the fragments in mass spectrometry. Incubation of gliadins withRothiacells reduced R5 antibody binding by 20, 82, and 97% after 30 min, 2 h, and 5 h, respectively, which was paralleled by reduced reactivity of enzyme-treated 33-mer and 26-mer peptides in the R5 competitive ELISA. Our broad complementary approach to validate gluten degrading activities qualifiesR. mucilaginosa-associated enzymes as promising tools to neutralize T cell immunogenic properties for treatment of celiac disease.
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DOI：
10.1097/01.meg.0000059119.41030.df
发表时间：
2003-05-01
影响因子：
2.1
作者：
Valdés, I;García, E;Méndez, E
通讯作者：
Méndez, E
组织转谷氨酰胺酶的选择性脱酰胺作用强烈增强麦醇溶蛋白特异性 T 细胞反应性。
影响因子：
4.4
作者：
Y. van de Wal;Y. Kooy;P. V. van Veelen;S. Peña;L. Mearin;G. Papadopoulos;F. Koning
通讯作者：
F. Koning
影响因子：
24.5
作者：
Cerf-Bensussan, Nadine;Matysiak-Budnik, Tamara;Heyman, Martine
通讯作者：
Heyman, Martine
影响因子：
4.4
作者：
Shan, L;Qiao, SW;Khosla, C
通讯作者：
Khosla, C
影响因子：
2.9
作者：
L. Lorand;L. Campbell
通讯作者：
L. Campbell