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Title:Analysis of the ammonium assimilation pathways of the human colonic bacterium, bacteroides thetaiotaomicron
Author(s):Iakiviak, Michael
Director of Research:Mackie, Roderick I.
Doctoral Committee Chair(s):Mackie, Roderick I.
Doctoral Committee Member(s):Cann, Isaac K.O.; Ridlon, Jason; Degnan, Patrick
Department / Program:Animal Sciences
Discipline:Animal Sciences
Degree Granting Institution:University of Illinois at Urbana-Champaign
Degree:Ph.D.
Genre:Dissertation
Subject(s):Bacteroides
Ammonium assimilation
Abstract:In ruminants, efficient rumen function and proper host metabolism is dependent on the nitrogen supply in the feed. Assimilated ammonium accounts for up to 70% of the microbial protein production, which satisfies up to 85% of the host protein requirements. Similar numbers for the human colon have not been determined. However, colonic bacteria are responsible for the production of ammonium, derived from host-secreted urea and endogenous and dietary proteins, that provides the preferred nitrogen source for microbial growth. Bacteroides thetaiotaomicron, a model organism for human gut Bacteroidetes, encodes genes for the capture of ammonium through the two primary pathways, the glutamate dehydrogenase (GDH) pathway and the glutamine synthetase/glutamate synthase (GS/GOGAT) pathway. To gain insight into the genomic features underlying ammonium uptake and assimilation in this bacterium, comparative transcriptomic analysis using RNA-Seq was employed on cultures growing under excess or limiting ammonium concentrations. A single genomic locus, encoding for the GS/GOGAT pathway, was identified with highly increased transcription when the organism grows under limiting ammonium concentration. The relative contribution of each gene to ammonium assimilation was assessed through construction of genomic deletion strains for each of the three GS, one GOGAT, and two GDH genes. The deletion of two genes, the NADPH-dependent glutamate dehydrogenase (gdhA) and the glutamine synthetase type 3 (glnN2) significantly impeded growth of the organisms under both nitrogen conditions. Taken together, the results demonstrate the importance of the GDH pathway for constitutive ammonium assimilation, and GlnN2 for glutamine biosynthesis. However, when the organism grows under nitrogen limitation, the GS/GOGAT pathway, including glnN1, is highly induced. To extrapolate the significance of the findings, a comparative bioinformatic analysis, using all of the available sequenced Bacteroides genomes, revealed high conservation of the critical genomic loci in gut species. Understanding of nitrogen metabolism in gut microbes is essential for a complete depiction of their ecological implications on the host´s metabolism in health and disease.
Issue Date:2017-12-06
Type:Text
URI:http://hdl.handle.net/2142/99511
Rights Information:Copyright 2017 Michael Iakiviak
Date Available in IDEALS:2018-03-13
2020-03-14
Date Deposited:2017-12


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