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Development of an enhanced metaproteomic approach for deepening the microbiome characterization of the human infant gut.

Xiong W, Giannone RJ, Morowitz MJ, Banfield JF, Hettich RL - J. Proteome Res. (2014)

Bottom Line: To alleviate this restriction, we have designed a novel metaproteomic strategy based on double filtering (DF) the raw samples, a method that fractionates microbial from human cells to enhance microbial protein identification and characterization in complex fecal samples from healthy premature infants.This method dramatically improved the overall depth of infant gut proteome measurement, with an increase in the number of identified low-abundance proteins and a greater than 2-fold improvement in microbial protein identification and quantification.This enhancement of proteome measurement depth enabled a more extensive microbiome comparison between infants by not only increasing the confidence of identified microbial functional categories but also revealing previously undetected categories.

View Article: PubMed Central - PubMed

Affiliation: Chemical Sciences Division, Oak Ridge National Laboratory , Oak Ridge, Tennessee 37831, United States.

ABSTRACT
The establishment of early life microbiota in the human infant gut is highly variable and plays a crucial role in host nutrient availability/uptake and maturation of immunity. Although high-performance mass spectrometry (MS)-based metaproteomics is a powerful method for the functional characterization of complex microbial communities, the acquisition of comprehensive metaproteomic information in human fecal samples is inhibited by the presence of abundant human proteins. To alleviate this restriction, we have designed a novel metaproteomic strategy based on double filtering (DF) the raw samples, a method that fractionates microbial from human cells to enhance microbial protein identification and characterization in complex fecal samples from healthy premature infants. This method dramatically improved the overall depth of infant gut proteome measurement, with an increase in the number of identified low-abundance proteins and a greater than 2-fold improvement in microbial protein identification and quantification. This enhancement of proteome measurement depth enabled a more extensive microbiome comparison between infants by not only increasing the confidence of identified microbial functional categories but also revealing previously undetected categories.

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COG categoryanalysis of microbial protein groups. Microbial proteingroups are assigned into COG categories via rpsblast against the COGdatabase from NCBI. Distributions of identified categories were constructedby category counts and spectra of infant #UN1 (a) and infant #CA1(b). Abundant categories are numerically labeled.
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fig5: COG categoryanalysis of microbial protein groups. Microbial proteingroups are assigned into COG categories via rpsblast against the COGdatabase from NCBI. Distributions of identified categories were constructedby category counts and spectra of infant #UN1 (a) and infant #CA1(b). Abundant categories are numerically labeled.

Mentions: To furtherelucidate the advantages of the DF sample preparation approach, wetabulated and analyzed the COG functions for the two infant gut microbiomesmeasured by these two methods (Table S5). We clearly recognize that COG families are relatively broad andcharacterize the functionality at a lower resolution than a more specific,detailed metabolic pathway analysis. Nevertheless, they were usedhere to provide only a general metric for the power of the microbialprotein enriching approach and not necessarily to assess biologicaldifferences between the two infants. For both infants, we found severalhighly represented COG categories, including carbohydrate transportand metabolism; energy production and conversion; translation, ribosomalstructure, and biogenesis; post-translational modification, proteinturnover, chaperones; and amino acid transport and metabolism (Figure 5). A similar distribution of COG functions was reportedfor a healthy adult twin pair.24 Theseresults suggest that the establishment of microbial communities inthese two infants gut environments is fairly quickly migrating towarda relatively stable and adult-like microbiota, which plays a crucialrole in carbohydrate metabolism and nutrient production.


Development of an enhanced metaproteomic approach for deepening the microbiome characterization of the human infant gut.

Xiong W, Giannone RJ, Morowitz MJ, Banfield JF, Hettich RL - J. Proteome Res. (2014)

COG categoryanalysis of microbial protein groups. Microbial proteingroups are assigned into COG categories via rpsblast against the COGdatabase from NCBI. Distributions of identified categories were constructedby category counts and spectra of infant #UN1 (a) and infant #CA1(b). Abundant categories are numerically labeled.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4286196&req=5

fig5: COG categoryanalysis of microbial protein groups. Microbial proteingroups are assigned into COG categories via rpsblast against the COGdatabase from NCBI. Distributions of identified categories were constructedby category counts and spectra of infant #UN1 (a) and infant #CA1(b). Abundant categories are numerically labeled.
Mentions: To furtherelucidate the advantages of the DF sample preparation approach, wetabulated and analyzed the COG functions for the two infant gut microbiomesmeasured by these two methods (Table S5). We clearly recognize that COG families are relatively broad andcharacterize the functionality at a lower resolution than a more specific,detailed metabolic pathway analysis. Nevertheless, they were usedhere to provide only a general metric for the power of the microbialprotein enriching approach and not necessarily to assess biologicaldifferences between the two infants. For both infants, we found severalhighly represented COG categories, including carbohydrate transportand metabolism; energy production and conversion; translation, ribosomalstructure, and biogenesis; post-translational modification, proteinturnover, chaperones; and amino acid transport and metabolism (Figure 5). A similar distribution of COG functions was reportedfor a healthy adult twin pair.24 Theseresults suggest that the establishment of microbial communities inthese two infants gut environments is fairly quickly migrating towarda relatively stable and adult-like microbiota, which plays a crucialrole in carbohydrate metabolism and nutrient production.

Bottom Line: To alleviate this restriction, we have designed a novel metaproteomic strategy based on double filtering (DF) the raw samples, a method that fractionates microbial from human cells to enhance microbial protein identification and characterization in complex fecal samples from healthy premature infants.This method dramatically improved the overall depth of infant gut proteome measurement, with an increase in the number of identified low-abundance proteins and a greater than 2-fold improvement in microbial protein identification and quantification.This enhancement of proteome measurement depth enabled a more extensive microbiome comparison between infants by not only increasing the confidence of identified microbial functional categories but also revealing previously undetected categories.

View Article: PubMed Central - PubMed

Affiliation: Chemical Sciences Division, Oak Ridge National Laboratory , Oak Ridge, Tennessee 37831, United States.

ABSTRACT
The establishment of early life microbiota in the human infant gut is highly variable and plays a crucial role in host nutrient availability/uptake and maturation of immunity. Although high-performance mass spectrometry (MS)-based metaproteomics is a powerful method for the functional characterization of complex microbial communities, the acquisition of comprehensive metaproteomic information in human fecal samples is inhibited by the presence of abundant human proteins. To alleviate this restriction, we have designed a novel metaproteomic strategy based on double filtering (DF) the raw samples, a method that fractionates microbial from human cells to enhance microbial protein identification and characterization in complex fecal samples from healthy premature infants. This method dramatically improved the overall depth of infant gut proteome measurement, with an increase in the number of identified low-abundance proteins and a greater than 2-fold improvement in microbial protein identification and quantification. This enhancement of proteome measurement depth enabled a more extensive microbiome comparison between infants by not only increasing the confidence of identified microbial functional categories but also revealing previously undetected categories.

Show MeSH