Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2013;8(2):e57535.
doi: 10.1371/journal.pone.0057535. Epub 2013 Feb 26.

Proteomic analysis of Bifidobacterium longum subsp. infantis reveals the metabolic insight on consumption of prebiotics and host glycans

Jae-Han Kim  1 , Hyun Joo An, Daniel Garrido, J Bruce German, Carlito B Lebrilla, David A Mills
Affiliations

Proteomic analysis of Bifidobacterium longum subsp. infantis reveals the metabolic insight on consumption of prebiotics and host glycans

Jae-Han Kim et al. PLoS One. 2013.

Abstract

Bifidobacterium longum subsp. infantis is a common member of the intestinal microbiota in breast-fed infants and capable of metabolizing human milk oligosaccharides (HMO). To investigate the bacterial response to different prebiotics, we analyzed both cell wall associated and whole cell proteins in B. infantis. Proteins were identified by LC-MS/MS followed by comparative proteomics to deduce the protein localization within the cell. Enzymes involved in the metabolism of lactose, glucose, galactooligosaccharides, fructooligosaccharides and HMO were constitutively expressed exhibiting less than two-fold change regardless of the sugar used. In contrast, enzymes in N-Acetylglucosamine and sucrose catabolism were induced by HMO and fructans, respectively. Galactose-metabolizing enzymes phosphoglucomutase, UDP-glucose 4-epimerase and UTP glucose-1-P uridylytransferase were expressed constitutively, while galactokinase and galactose-1-phosphate uridylyltransferase, increased their expression three fold when HMO and lactose were used as substrates for cell growth. Cell wall-associated proteomics also revealed ATP-dependent sugar transport systems associated with consumption of different prebiotics. In addition, the expression of 16 glycosyl hydrolases revealed the complete metabolic route for each substrate. Mucin, which possesses O-glycans that are structurally similar to HMO did not induced the expression of transport proteins, hydrolysis or sugar metabolic pathway indicating B. infantis do not utilize these glycoconjugates.

PubMed Disclaimer

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. Distribution of the NSAF ratio of each SBP between soluble and insoluble fraction (grey bars; ψNSAFinsol/NSAFsol).
White bars (soluble: left corner) and black bars (insoluble: right corner) indicate the number of SBP observed solely in soluble and insoluble fraction, respectively.
Figure 2
Figure 2. Central metabolic pathways reconstructed from proteomic datasets.
Sugar substrates are presented inside the red boxes. Pathways regulated or induced by the presence of a specific sugar are indicated by colored background and their expressions on different sugars are presented next to the pathway as bar graphs: (a) Leloir pathway, (b) HexNAc catabolic pathway, (c) pyruvate fermentation pathway, (d) FOS/Inulin glycosyl hydrolase. The relative amounts of enzymes involved in the bifid shunt are presented in Figure S3. Numbers in the parenthesis next to the enzyme are the locus tag of each protein expressed.
Figure 3
Figure 3. The expression of Family 1 extracellular SBPs on different carbon sources.
The quantitative expression of each SBP is described by NSAF value. Black and gray bars indicate the NSAF values obtained in insoluble and soluble fraction, respectively. Box indicates the SBPs in the HMO cluster.
Figure 4
Figure 4. The expression of proteins in HMO cluster in B. infantis during growth on different prebiotics.
(a) Graphical display of genes in HMO cluster. Proteins involved in the HMO translocation and degradation are noted with the locus tag under the gene. Sets of genes inside the red box are the transport systems that comprise a Family 1 extracellular SBP and two inner-membrane components. Bars represent the relative amounts of each protein in different carbon sources. Black and gray bars indicate the NSAF values in the soluble and insoluble fraction, respectively. (b)∼(e) are the glycosyl hydrolases for HMO degradation. (f)∼(k) are the Family 1 SBPs and (l)∼(n) are the proteins whose the amount was high but the role in HMO metabolism is not well known.
Figure 5
Figure 5. The expression of glycosyl hydrolases and glycosyltransferases in B. infantis during growth on different prebiotics.
The amount of protein expressed scaled by color from green to red as shown in the legend. Gray indicates that the expression was not detected. Left and right panel of heat map represent the expression of protein in soluble and insoluble fraction, respectively. CWA protein was determined by the NSAF ratio between soluble and insoluble faction (NSAFinsol/NSAFsol≥2.5).
Figure 6
Figure 6. Schematic diagram of the potential metabolic pathways for different prebiotic consumption in B. infantis.
The catabolic pathways for monosaccharides were detailed in Figure 2. Bold and curved arrows indicate the flow of oligosaccharide.
See this image and copyright information in PMC

References

    1. Roger LC, McCartney AL (2010) Longitudinal investigation of the faecal microbiota of healthy full-term infants using fluorescence in situ hybridization and denaturing gradient gel electrophoresis. Microbiology 156: 3317–3328. - PubMed
    1. Yatsunenko T, Rey FE, Manary MJ, Trehan I, Dominguez-Bello MG, et al. (2012) Human gut microbiome viewed across age and geography. Nature 486: 222–227. - PMC - PubMed
    1. Ninonuevo MR, Ward RE, LoCascio RG, German JB, Freeman SL, et al. (2007) Methods for the quantitation of human milk oligosaccharides in bacterial fermentation by mass spectrometry. Analytical Biochemistry 361: 15–23. - PubMed
    1. Tao N, Depeters EJ, Freeman S, German JB, Grimm R, et al. (2008) Bovine milk glycome. Journal of Dairy Science 91: 3768–3778. - PubMed
    1. Ninonuevo MR, Park Y, Yin H, Zhang J, Ward RE, et al. (2006) A strategy for annotating the human milk glycome. J Agric Food Chem 54: 7471–7480. - PubMed

Publication types