Protein composition and methods for analysing microbiota
Inventors
FIGEYS, Joseph Michel Daniel • STINTZI, Alain Christophe • Mack, David R. • Zhang, Xu • NING, Zhibin
Assignees
Publication Number
US-11933790-B2
Publication Date
2024-03-19
Expiration Date
2037-06-01
Interested in licensing this patent?
MTEC can help explore whether this patent might be available for licensing for your application.
Abstract
A method of isotope-labelling a microbiota sample. It involves providing a first microbiota sample that was obtained from a given source; exposing the first microbiota sample to an isotope enriched medium; and culturing the exposed first microbiota sample in the isotope enriched medium to obtain an isotope-labelled microbiota sample, wherein the isotope labelled metaproteome of the isotope-labelled microbiota sample is taxon specific for taxa present in the first microbiota sample when initially obtained from the given source.
Core Innovation
The invention provides a method and composition for isotope-labelling a microbiota sample by exposing the microbiota sample, obtained from a given source, to an isotope enriched medium and culturing it to obtain an isotope-labelled microbiota sample with a taxon-specific labelled metaproteome representative of taxa present originally. This method, termed stable isotopically labelled microbiota (SILAMi), allows for metabolic labelling of complex microbiota including diverse bacteria, archaea, and eukaryotes across different microbiota types from humans, animals, soil, plants, or other environments.
The problem solved by the invention arises from the challenges in metaproteomics of microbiota samples due to high species diversity and difficulty in culturing diverse anaerobic and aerobic microbes. Existing methods like label free quantification (LFQ) suffer from high variability and metabolic labelling methods such as SILAC and SILAM have limited application to complex microbiota. There is a need for a reliable, consistent, and representative isotope-labelled standard for the whole microbiota to improve identification and quantification of microbial peptides/proteins, providing more accurate functional insight into microbiota composition and response to various compounds.
The invention overcomes previous assumptions that diverse microbiota populations cannot be labelled simultaneously or reliably due to changes in microbial composition during incorporation of the labelling isotope. SILAMi provides a fast, cost-effective, and efficient approach to achieve highly enriched isotopic labelling (over 90% average heavy isotope enrichment), maintaining taxon-specific representation of the microbiota sample. The labelled standard serves as an internal standard for metaproteomic analysis, enabling accurate quantification, reduction of experimental variability, and assessment of microbiota composition and functionality, including changes induced by treatment or environmental exposures.
Claims Coverage
The patent contains multiple inventive features focusing on a method for high throughput screening of microbiota samples using isotope-labelled standards and metaproteomic analysis.
Method of high throughput screening of microbiota samples using isotope-labelled standards
Providing proteome extracts from multiple microbiota samples from humans, soil, or animals cultured and spiking them with an isotope-labelled microbiota proteome standard obtained by culturing microbiota in isotope enriched medium, followed by pre-screening via metaproteomic analysis, selecting microbiomes with changes, and analyzing these microbiomes to characterize the changes.
Use of microbial gene catalog and iterative database search for analysis
Analyzing selected microbiomes with a microbial gene catalog of a given subject type (human or animal) and using an iterative database search strategy to improve characterization of microbiome changes.
Combination of metaproteomic and metagenomic analysis
Performing metaproteomic analysis combined with metagenomic analysis for comprehensive assessment of microbiome changes.
Spiking labeled standard at a defined protein ratio
Adding sufficient isotope-labelled standard to proteome extracts to reach a 1:1 protein mass ratio with the native proteins in the microbiota samples to enable accurate quantification.
Application of analysis results to diagnostics and screening
Assessing analysis results for applications such as disease diagnosis, treatment response evaluation, remission assessment, and screening of xenobiotic or compound effects including food, drugs, chemicals, cosmetics, packaging materials, pesticides, and consumer products on microbiomes, and identifying subject responsiveness to therapies or treatments.
High enrichment and taxon specificity in the isotope-labelled standard
The isotope-labelled standard comprises proteins representative of the microbiota metaproteome with at least 90% average heavy isotopic enrichment rate and taxon-specific labelling for at least about 50% to 85% or more of the microbe populations present in the samples from humans, soil, or animals.
Use across diverse microbiota types and isotope types
Applicability of the method for microbiota samples from various human body sites including intestinal, vaginal, oral, cutis, bladder, kidney, lung, eye, breast, penile microbiota, and use of stable or radioactive isotopes such as 13C, 14C, 15N, 32S, 35S, 32P, or deuterium or combinations thereof for labelling.
The claims cover an advanced method of microbiota metaproteomic analysis using isotope-labelled standards generated by culturing microbiota in isotope enriched media. The features emphasize high throughput screening, accuracy via taxon-specific labelling and isotopic enrichment, combined multi-omic analyses, and broad applications in diagnostics, treatment assessment, and compound screening across diverse microbiota sources and isotopes.
Stated Advantages
Provides a fast, efficient, and cost-effective method to generate metabolically labelled proteomes of complex microbiota with high isotope enrichment (over 90%).
Enables accurate, standardized, and reproducible quantitative metaproteomic analysis of microbiota samples, reducing inter- and intra-experiment variability.
Allows for improved identification and quantification of microbial proteins, including taxon-specific labelling representative of the original microbiota population.
Facilitates detection and analysis of microbiota functional changes in response to compounds, disease state, treatment response, or environmental exposures.
Supports high throughput screening of multiple microbiota samples for rapid classification of compound effects on microbiota composition and function.
Documented Applications
Screening and assessing effects of xenobiotics, drugs, chemicals, food compounds, beverages, food additives, cosmetics, cosmetic components, packaging materials, pesticides, herbicides, consumer products, and therapeutic agents on human, animal, soil, and biofilm microbiota.
Diagnostic use including disease diagnosis, assessing treatment response, and determining remission status by analyzing microbiota protein expression changes.
Rapid screening of individual or multiple microbiome samples using RapidAIM platform in multi-well formats for metaproteomic and metagenomic functional analysis.
Use of isotope-labelled microbiota protein standards to improve accuracy of metaproteomic quantification and functional interpretation in diverse microbiota samples.
Interested in licensing this patent?