Treatment of gluten intolerance and related conditions

A technology for gluten intolerance and related diseases, which is applied in the field of compositions for treating gluten intolerance and related diseases, and can solve problems such as bacteria species that are difficult to eradicate

Active Publication Date: 2016-04-13
CODEXIS INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In addition, some bacterial species form endospores

Method used

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  • Treatment of gluten intolerance and related conditions
  • Treatment of gluten intolerance and related conditions
  • Treatment of gluten intolerance and related conditions

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0157] Preparation of Nepenthes Protease Extract

[0158] Chemicals

[0159] HPLC grade water and acetonitrile from Burdick and Jackson were purchased from VWR. Formic acid, Tris, glycine were purchased from SigmaAldrich.

[0160] plant culture

[0161] Transplants of Nepenthes rafflesiana, Nepenthes ampularia, Nepenthes mirabilis and Nepenthes globosa were purchased from Keehns Carnivores (http: / / www.keehnscarnivores.ca) . These transplanted plants were cultivated with veneer, perlite, peat moss and humus (40%, 35%, 10%, 5%, respectively). The growing conditions include 14 hours of light per day, 80% humidity and a temperature of 23°C to 28°C and watering two to three times a week. After the pitcher leaves of Nepenthes matured, the plants were fed with one or two flies per pitcher, and pitcher sap was harvested one week later. The recovery of pitcher leaves and their secretions lasted one week, followed by a second round of feeding and extraction.

[0162] Extract prep...

Embodiment 2

[0167] Protein Digestion Mapping of Pepsin and Nepenthes Protease Extracts

[0168] Digestion mapping of nepenthes proteases by mass spectrometry

[0169] Nepenthes protease extract was prepared as in Example 1.

[0170] Protein digestion was performed in solution using a LEAPHTX-PAL autosampler and a distribution system designed for hydrogen / deuterium exchange (HDX) applications, and data was collected using an ABSciex Triple-TOF5600QqTOF mass spectrometer. Peptides were identified by MS / MS data using Mascot (v2.3). Briefly, 8 μL, 8 μM protein (XRCC4, XLF, Ligase IV-tandem BRCT domain, PNK, myoglobin or cytochrome C) was mixed with 10 μL, 11-fold concentrated liquid for 2 min at 10°C. Myoglobin and cytochrome C were purchased from Sigma. After dilution to 1 μM substrate concentration, 15 μL was injected into a refrigerated reverse phase LC system (4° C.) connected to a mass spectrometer. Peptides were captured on a 5 cm, 200 μm.d. Onyx C18 monolithic column (Phenomenex In...

Embodiment 3

[0175] Digestion mapping of the multidomain protein XRCC4 by pepsin and nepenthes protease extracts

[0176] HD exchange of complexes involved in DNA damage repair

[0177] Nepenthes protease extract was prepared as in Example 1.

[0178] Stock solutions of XRCC4 (1-200) containing BRCT as well as XRCC4 (full length) containing BRCT were diluted to equimolar concentration (10 μM) in buffer (10 mM Tris-HCl, pH 7.5) and incubated at 4°C for minimal 30 minutes for enhanced complexation. Samples were kept at 4°C until HDX analysis. Aliquot the sample at 20 °C by adding D 2O (25% v / v) deuterated for 2 minutes. Aliquots were subsequently digested in two ways. In the first digestion strategy, protein deuteration was quenched by adding the sample to chilled 100 mM glycine-HCl, pH 2.5, and the quenched protein solution was injected into a pepsin microreactor. The microreactor was installed in the HTX-PAL system between the injector valve and the C18 column. Protein digests were ...

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Abstract

Provided herein are compositions, foods comprising nepenthesin or a derivative thereof and methods of using nepenthesin or a derivative thereof for modulating gluten intolerance and related conditions, such as celiac disease. Further provided herein are pharmaceutical compositions comprising nepenthesin or a derivative thereof and methods of using nepenthesin or a derivative thereof to treat bacterial infections of the gastrointestinal tract, such as C. difficile or H. pylori. Further provided herein are compositions comprising recombinant nepenthesin I or nepenthesin II, or homologous proteins, and methods for making the same.

Description

technical field [0001] Provided herein are compositions, foods, and methods for treating gluten intolerance and related conditions (eg, celiac disease). Further provided herein are pharmaceutical compositions comprising nepenthesin or derivatives thereof and methods of using nepenthesin or derivatives thereof to treat gastrointestinal bacterial (eg, Clostridium difficile or H. pylori) infections. Also provided herein are methods of using nepenthesin or derivatives thereof in hydrogen / deuterium exchange. Further provided herein are compositions comprising recombinant nepenthesin I or nepenthesin II or homologous proteins and methods for their preparation. Background technique [0002] In individuals with gluten intolerance, digestion of gluten-containing wheat, barley, rye, and possibly oats may cause abnormal autoimmune reactions, eg, celiac disease, wheat allergy, and dermatitis herpetiformis. Gluten is a mixture of glutenin and gliadin molecules rich in glutamine and pro...

Claims

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Application Information

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IPC IPC(8): C12N9/50A23L33/00A61K38/48A61P1/14A61P31/04A23L29/00
CPCA23V2002/00C12Y304/23012A23L29/06A23L33/10A61K38/488A23V2200/304A23V2250/5486
Inventor 戴维·施里默彼得·曼海尼克·马策克马尔什·雷伊
Owner CODEXIS INC
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