Identification of food-grade subtilisins as gluten-degrading enzymes to treat celiac disease.
Identification of food-grade subtilisins as gluten-degrading enzymes to treat celiac disease.
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鉴定食品级枯草杆菌蛋白酶作为治疗乳糜泻的麸质降解酶。
DOI：
10.1152/ajpgi.00185.2016
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发表时间：
2016
期刊：
影响因子：
--
通讯作者：
Helmerhorst,EvaJ
中科院分区：
文献类型：
--
作者：
Wei,Guoxian;Tian,Na;Siezen,Roland;Schuppan,Detlef;Helmerhorst,EvaJ
谷蛋白是富含脯氨酸和谷氨酰胺的蛋白质，存在于小麦、大麦和黑麦中，含有导致乳糜泻（CD）的免疫原性序列。粘胶罗氏菌，一种口腔微生物定植剂，可以切割这些谷蛋白表位。目的是分离和鉴定这些酶，并评估它们作为治疗乳糜泻的新酶的潜力。用DEAE层析法提取并分离黏液蛋白。利用副硝基苯胺衍生物和荧光共振能量转移（FRET）肽底物以及麦胶蛋白酶谱法监测酶活性。R5和G12 elisa检测表位消除。通过LC-ESI-MS/MS鉴定了三种降解麦胶蛋白的rothia酶，分别为ROTMU0001_0241 （C6R5V9_9MICC）、ROTMU0001_0243 （C6R5W1_9MICC）和ROTMU0001_240 （C6R5V8_9MICC）。使用Basic Local Alignment search Tool进行搜索显示，这些是属于肽酶S8家族的枯草杆菌样丝氨酸蛋白酶。对主要的rothiasubtilisins进行比对表明，它们都含有Asp (D)、His (H)和Ser (S)按D-H-S顺序的催化三联体。他们裂解了枯草菌素的底物琥珀酰- ala - ala -Pro- ph -副硝基苯胺，其中Pro位于P2位置，就像酪氨酸-Pro- gln和酪氨酸-亮氨酸-Pro- tyr一样，它们也被裂解。一致地，含有Xaa-Pro-Xaa动机的麦胶蛋白免疫原性表位的FRET底物被快速水解。地衣芽孢杆菌中的枯草素A和食品级纳豆激酶能有效降解免疫原性麦胶蛋白衍生的33-mer肽和R5和G12抗体识别的免疫显性表位。本研究确定罗氏和食品级的枯草芽孢杆菌是有前途的新候选酶治疗乳糜泻。
Gluten are proline- and glutamine-rich proteins present in wheat, barley, and rye and contain the immunogenic sequences that drive celiac disease (CD).Rothia mucilaginosa, an oral microbial colonizer, can cleave these gluten epitopes. The aim was to isolate and identify the enzymes and evaluate their potential as novel enzyme therapeutics for CD. The membrane-associatedR. mucilaginosaproteins were extracted and separated by DEAE chromatography. Enzyme activities were monitored with paranitroanilide-derivatized and fluorescence resonance energy transfer (FRET) peptide substrates, and by gliadin zymography. Epitope elimination was determined in R5 and G12 ELISAs. The gliadin-degradingRothiaenzymes were identified by LC-ESI-MS/MS as hypothetical proteins ROTMU0001_0241 (C6R5V9_9MICC), ROTMU0001_0243 (C6R5W1_9MICC), and ROTMU0001_240 (C6R5V8_9MICC). A search with the Basic Local Alignment Search Tool revealed that these are subtilisin-like serine proteases belonging to the peptidase S8 family. Alignment of the majorRothiasubtilisins indicated that all contain the catalytic triad with Asp (D), His (H), and Ser (S) in the D-H-S order. They cleaved succinyl-Ala-Ala-Pro-Phe-paranitroanilide, a substrate for subtilisin with Pro in the P2 position, as in Tyr-Pro-Gln and Leu-Pro-Tyr in gluten, which are also cleaved. Consistently, FRET substrates of gliadin immunogenic epitopes comprising Xaa-Pro-Xaa motives were rapidly hydrolyzed. TheRothiasubtilisins and two subtilisins fromBacillus licheniformis, subtilisin A and the food-grade Nattokinase, efficiently degraded the immunogenic gliadin-derived 33-mer peptide and the immunodominant epitopes recognized by the R5 and G12 antibodies. This study identifiedRothiaand food-gradeBacillussubtilisins as promising new candidates for enzyme therapeutics in CD.
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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：
3.2
作者：
Branchi, Federica;Tomba, Carolina;Elli, Luca
通讯作者：
Elli, Luca
影响因子：
2.9
作者：
Siezen, Roland J.;Renckens, Bernadet;Boekhorst, Jos
通讯作者：
Boekhorst, Jos
枯草杆菌蛋白酶抑制剂eglin c中的活性位点结合环稳定性：对与枯草杆菌蛋白酶和未复合的抑制剂复合的专门设计的突变体进行结构和功能研究。
影响因子：
--
作者：
K. Hipler;J. Priestle;J. Rahuel;M. Grütter
通讯作者：
M. Grütter