Smirnov K.S.,Helmholtz Center Munich |
Maier T.V.,Helmholtz Center Munich |
Walker A.,Helmholtz Center Munich |
Heinzmann S.S.,Helmholtz Center Munich |
And 6 more authors.
International Journal of Medical Microbiology | Year: 2016
The review highlights the role of metabolomics in studying human gut microbial metabolism. Microbial communities in our gut exert a multitude of functions with huge impact on human health and disease. Within the meta-omics discipline, gut microbiome is studied by (meta)genomics, (meta)transcriptomics, (meta)proteomics and metabolomics. The goal of metabolomics research applied to fecal samples is to perform their metabolic profiling, to quantify compounds and classes of interest, to characterize small molecules produced by gut microbes. Nuclear magnetic resonance spectroscopy and mass spectrometry are main technologies that are applied in fecal metabolomics. Metabolomics studies have been increasingly used in gut microbiota related research regarding health and disease with main focus on understanding inflammatory bowel diseases. The elucidated metabolites in this field are summarized in this review. We also addressed the main challenges of metabolomics in current and future gut microbiota research. The first challenge reflects the need of adequate analytical tools and pipelines, including sample handling, selection of appropriate equipment, and statistical evaluation to enable meaningful biological interpretation. The second challenge is related to the choice of the right animal model for studies on gut microbiota. We exemplified this using NMR spectroscopy for the investigation of cross-species comparison of fecal metabolite profiles. Finally, we present the problem of variability of human gut microbiota and metabolome that has important consequences on the concepts of personalized nutrition and medicine. © 2016 Elsevier GmbH.
Stelzl T.,TU Munich |
Stelzl T.,Institute for Food and Health |
Baranov T.,TU Munich |
Baranov T.,Institute for Food and Health |
And 6 more authors.
American Journal of Physiology - Gastrointestinal and Liver Physiology | Year: 2016
The intestinal peptide transporter PEPT1 provides bulk quantities of amino acids to epithelial cells. PEPT1 is a high-capacity and lowaffinity solute carrier of the SLC15 family found in apical membranes of enterocytes in small intestine and distal colon. Surprisingly, murine PEPT1 (mPEPT1) has an apparent molecular mass of ~95 kDa in the small intestine but ~105 kDa in the large intestine. Here we describe studies on mPEPT1 protein glycosylation and how glycans affect transport function. Putative N-glycosylation sites of mPEPT1 were altered by site-directed mutagenesis followed by expression in Xenopus laevis oocytes. Replacement of six asparagine residues (N) at positions N50, N406, N439, N510, N515, and N532 by glutamine (Q) resulted in a decrease of the mPEPT1 mass by around 35 kDa. Electrophysiology revealed all glycosylation-deficient transporters to be functional with comparable expression levels in oocyte membranes. Strikingly, the mutant protein with N50Q exhibited a twofold decreased affinity for Gly-Sar but a 2.5-fold rise in the maximal inward currents compared with the wild-type protein. Elevated maximal transport currents were also recorded for cefadroxil and tri-Lalanine. Tracer flux studies performed with [14C]-Gly-Sar confirmed the reduction in substrate affinity and showed twofold increased maximal transport rates for the N50Q transporter. Elimination of individual N-glycosylation sites did not alter membrane expression in oocytes or overall transport characteristics except for the mutant protein N50Q. Because transporter surface density was not altered in N50Q, removal of the glycan at this location appears to accelerate the substrate turnover rate. © 2016 the American Physiological Society.