The NIH Human Microbiome Project (HMP) revolutionized our perspective on human-microbe interactions and provided a tremendous impetus for research in order to obtain a much deeper understanding of how microbes impact human health. The gut microorganisms of the HMP Reference Genomes and the Human Gastrointestinal Bacteria Culture Collection contain 961 species representing 142 genera. Yet, relatively little is known about these microorganisms. Herein we will test the hypothesis that tungsten (W), a metal almost never considered in biological systems, is essential for the health of the human gut microbiome. Our bioinformatics analyses reveal that a large number of these gut microbes contain genes encoding diverse members of the W-containing oxidoreductase (WOR) family of enzymes. Only a very few WOR enzymes have been previously characterized, mainly from exotic thermophilic microbes. The overall goal of the proposed research is to show that other members of the WOR family have essential functions in the gut microbiome. In preliminary studies, we have shown that some gut microbes take up trace amounts of W and their W-containing WORs remove reactive and potentially toxic aldehydes found in the gut, which are generated from cooked foods and microbiome metabolism. Other W-containing WORs are proposed to catalyze other as yet unknown reactions. In the proposed research we will purify ten novel phylogenetically distinct WORs by W-monitored (using ICP-MS) anaerobic chromatography. Their catalytic activities and physiological substrates will be determined by an enzyme-induced metabolomics approach (using LC-MS). In addition, we propose that some of these WORs are electron bifurcating enzymes that simultaneously couple exergonic and endergonic reactions, a recently discovered mechanism of energy conservation in biological systems. Kinetic, spectroscopic (using EPR) and structural (using cryoEM) analyses of this subset of W-enzymes will be used to investigate the nature of the bifurcation reactions. Using genome-based metabolic reconstructions, the physiological functions of the various WORs will be ascertained and we will determine the effects of W on the metabolism of the gut microbes, including on their resistance to gut- and cooking-related aldehydes. Relevance: It is now clear that, in addition to facilitating digestion, the gut microbiome plays roles in a surprisingly extensive range of human conditions, including in Parkinson?s, schizophrenia, osteoarthritis and in cardiovascular and immune-deficiency diseases. The results of the proposed research will provide a completely new perspective on the primary metabolisms of the key microbes in our gastrointestinal tract and the proposed essential role of tungsten, a metal that was thought to be seldom used in biological systems.
The NIH human microbiome project lists over 800 reference microorganisms derived from the human gastrointestinal tract but relatively little is known about them and fundamental studies at the metabolic and molecular level are sorely needed. Herein we will test the hypothesis that tungsten (W), a metal almost never considered in biological systems, is essential for the health of the human gut microbiome. The overall goal of this research is to purify ten new types of tungstoenzyme from representative gut microbes grown in the laboratory and characterize them to determine their physiological and metabolic roles.
