Khushboo Borah

Dr Khushboo Borah B.Tech, DPhil


Research Fellow
Bachelor of Technology (engineering), DPhil

Biography

Areas of specialism

Gas chromatography mass spectrometry (GC-MS) and Liquid chromatography mass spectrometry (LC-MS/MS); Metabolomics; Tuberculosis; Leprosy; 13C Metabolic Flux Analysis; Oxysterol metabolism; 13C, 15N, 2H isotopic labelling analysis

University roles and responsibilities

  • Vice Chair Doctoral College Conference 2021

    My qualifications

    2013
    Bachelor of Technology (B.Tech) Biotechnology
    Indian Institute of Technology Guwahati India
    2018
    DPhil (Doctor of Philosophy) Plant Sciences
    University of Oxford
    2020
    Fellow of HEA
    University of Surrey

    Affiliations and memberships

    Research

    Research interests

    Research projects

    My publications

    Highlights

    • Metabolic Flux Analysis and Constraint based modelling to reveal metabolic control in rhizobia. Science Advances 2021
    • Metabolic fluxes for nutritional flexibility of Mycobacterium tuberculosis, Molecular Systems Biology 2021
    • Immune cell metabolism: Oxysterol metabolism is compartment specific. Redox Biology 2020
    • Tuberculosis: Mycobacterium tuberculosis uses multiple host nitrogen sources. Cell Reports 2019
    • Leprosy: Mycobacterium leprae uses host cell glucose as a carbon source. mBio 2019

    Publications

    Irundika Hk Dias, Khushboo Borah, Berivan Amin, Helen R Griffiths, Khouloud Sassi, Gérard Lizard, Ane Iriondo, Pablo Martinez-Lage (2019). Localisation of oxysterols at the sub-cellular level and in biological fluids
    View abstract View full publication
    Oxysterols are oxidized derivatives of cholesterol that are formed enzymatically or via reactive oxygen species or both. Cholesterol or oxysterols ingested as food are absorbed and packed into lipoproteins that are taken up by hepatic cells. Within hepatic cells, excess cholesterol is metabolised to form bile acids. The endoplasmic reticulum acts as the main organelle in the bile acid synthesis pathway. Metabolised sterols originating from this pathway are distributed within other organelles and in the cell membrane. The alterations to membrane oxysterol:sterol ratio affects the integrity of the cell membrane. The presence of oxysterols changes membrane fluidity and receptor orientation. It is well documented that hydroxylase enzymes located in mitochondria facilitate oxysterol production via an acidic pathway. More recently, the presence of oxysterols was also reported in lysosomes. Peroxisomal deficiencies favour intracellular oxysterols accumulation. Despite the low abundance of oxysterols compared to cholesterol, the biological actions of oxysterols are numerous and important. Oxysterol levels are implicated in the pathogenesis of multiple diseases ranging from chronic inflammatory diseases (atherosclerosis, Alzheimer's disease and bowel disease), cancer and numerous neurodegenerative diseases. In this article, we review the distribution of oxysterols in sub-cellular organelles and in biological fluids.
    Khushboo Borah, Martin Beyß, Axel Theorell, Huihai Wu, Piyali Basu, Tom A Mendum, Katharina Nӧh, Dany J V Beste, Johnjoe McFadden (2019). Intracellular Mycobacterium tuberculosis Exploits Multiple Host Nitrogen Sources during Growth in Human Macrophages
    View abstract View full publication
    Nitrogen metabolism of Mycobacterium tuberculosis (Mtb) is crucial for the survival of this important pathogen in its primary human host cell, the macrophage, but little is known about the source(s) and their assimilation within this intracellular niche. Here, we have developed 15N-flux spectral ratio analysis (15N-FSRA) to explore Mtb's nitrogen metabolism; we demonstrate that intracellular Mtb has access to multiple amino acids in the macrophage, including glutamate, glutamine, aspartate, alanine, glycine, and valine; and we identify glutamine as the predominant nitrogen donor. Each nitrogen source is uniquely assimilated into specific amino acid pools, indicating compartmentalized metabolism during intracellular growth. We have discovered that serine is not available to intracellular Mtb, and we show that a serine auxotroph is attenuated in macrophages. This work provides a systems-based tool for exploring the nitrogen metabolism of intracellular pathogens and highlights the enzyme phosphoserine transaminase as an attractive target for the development of novel anti-tuberculosis therapies.
    Khushboo Borah, Karina do Carmo de Vasconcelos Girardi, Tom A. Mendum, Leticia Miranda Santos Lery, Dany J. V. Beste, Flavio Alves Lara, Maria Cristina Vidal Pessolani, Johnjoe McFadden (2019). Intracellular Mycobacterium leprae Utilizes Host Glucose as a Carbon Source in Schwann Cellsdoi: 10.1128/mBio.02351-19
    View abstract View full publication
    New approaches are needed to control leprosy, but understanding of the biology of the causative agent Mycobacterium leprae remains rudimentary, principally because the pathogen cannot be grown in axenic culture. Here, we applied 13C isotopomer analysis to measure carbon metabolism of M. leprae in its primary host cell, the Schwann cell. We compared the results of this analysis with those of a related pathogen, Mycobacterium tuberculosis, growing in its primary host cell, the macrophage. Using 13C isotopomer analysis with glucose as the tracer, we show that whereas M. tuberculosis imports most of its amino acids directly from the host macrophage, M. leprae utilizes host glucose pools as the carbon source to biosynthesize the majority of its amino acids. Our analysis highlights the anaplerotic enzyme phosphoenolpyruvate carboxylase required for this intracellular diet of M. leprae, identifying this enzyme as a potential antileprosy drug target.
    Khushboo Borah, Jacque-Lucca Kearney, Ruma Banerjee, Pankaj Vats, Huihai Wu, Sonal Dahale, Sunitha Manjari Kasibhatla, Rajendra Joshi, Bhushan Bonde, Olabisi Ojo, Ramanuj Lahiri, Diana L Williams, Johnjoe McFadden (2020). GSMN-ML- a genome scale metabolic network reconstruction of the obligate human pathogen Mycobacterium leprae
    View abstract View full publication
    Leprosy, caused by Mycobacterium leprae, has plagued humanity for thousands of years and continues to cause morbidity, disability and stigmatization in two to three million people today. Although effective treatment is available, the disease incidence has remained approximately constant for decades so new approaches, such as vaccine or new drugs, are urgently needed for control. Research is however hampered by the pathogen's obligate intracellular lifestyle and the fact that it has never been grown in vitro. Consequently, despite the availability of its complete genome sequence, fundamental questions regarding the biology of the pathogen, such as its metabolism, remain largely unexplored. In order to explore the metabolism of the leprosy bacillus with a long-term aim of developing a medium to grow the pathogen in vitro, we reconstructed an in silico genome scale metabolic model of the bacillus, GSMN-ML. The model was used to explore the growth and biomass production capabilities of the pathogen with a range of nutrient sources, such as amino acids, glucose, glycerol and metabolic intermediates. We also used the model to analyze RNA-seq data from M. leprae grown in mouse foot pads, and performed Differential Producibility Analysis to identify metabolic pathways that appear to be active during intracellular growth of the pathogen, which included pathways for central carbon metabolism, co-factor, lipids, amino acids, nucleotides and cell wall synthesis. The GSMN-ML model is thereby a useful in silico tool that can be used to explore the metabolism of the leprosy bacillus, analyze functional genomic experimental data, generate predictions of nutrients required for growth of the bacillus in vitro and identify novel drug targets.
    Khushboo Borah, Olivia J. Rickman, Nikol Voutsina, Isaac Ampong, Dan Gao, Emma L. Baple, Irundika HK. Dias Andrew H. Crosby, Helen R. Griffiths (2020). A quantitative LC-MS/MS method for analysis of mitochondrial -specific oxysterol metabolism
    View abstract View full publication
    Oxysterols are critical regulators of inflammation and cholesterol metabolism in cells. They are oxidation products of cholesterol and may be differentially metabolised in subcellular compartments and in biological fluids. New analytical methods are needed to improve our understanding of oxysterol trafficking and the molecular interplay between the cellular compartments required to maintain cholesterol/oxysterol homeostasis. Here we describe a method for isolation of oxysterols using solid phase extraction and quantification by liquid chromatography-mass spectrometry, applied to tissue, cells and mitochondria. We analysed five monohydroxysterols; 24(S)-hydroxycholesterol, 25-hydroxycholesterol, 27-hydroxycholesterol, 7α-hydroxycholesterol, 7 ketocholesterol and three dihydroxysterols 7α-24(S)dihydroxycholesterol, 7α-25dihydroxycholesterol, 7α-27dihydroxycholesterol by LC-MS/MS following reverse phase chromatography. Our new method, using Triton and DMSO extraction, shows improved extraction efficiency and recovery of oxysterols from cellular matrix. We validated our method by reproducibly measuring oxysterols in mouse brain tissue and showed that mice fed a high fat diet had significantly lower levels of 24S/25diOHC, 27diOHC and 7ketoOHC. We measured oxysterols in mitochondria from peripheral blood mononuclear cells and highlight the importance of rapid cell isolation to minimise effects of handling and storage conditions on oxysterol composition in clinical samples. In addition,  cell culture systems, of THP-1 monocytes and neuronal-like SH-SH5Y cells, showed mitochondrial-specific oxysterol metabolism and profiles were lineage specific. In summary, we describe a robust and reproducible method validated for improved recovery, quantitative linearity and detection, reproducibility and selectivity for cellular oxysterol analysis. This method enables subcellular oxysterol metabolism to be monitored and is versatile in its application to various biological and clinical samples.
    in vitro
    Mackenzie JS, Lamprecht DA, Asmal R, Adamson JH, Borah K, Beste DJV, Lee BS, Pethe K, Rousseau S, Krieger I, Sacchettini JC, Glasgow JN, Steyn AJC (2020). Bedaquiline reprograms central metabolism to reveal glycolytic vulnerability in Mycobacterium tuberculosis
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    The approval of bedaquiline (BDQ) for the treatment of tuberculosis has generated substantial interest in inhibiting energy metabolism as a therapeutic paradigm. However, it is not known precisely how BDQ triggers cell death in Mycobacterium tuberculosis (Mtb). Using 13C isotopomer analysis, we show that BDQ-treated Mtb redirects central carbon metabolism to induce a metabolically vulnerable state susceptible to genetic disruption of glycolysis and gluconeogenesis. Metabolic flux profiles indicate that BDQ-treated Mtb is dependent on glycolysis for ATP production, operates a bifurcated TCA cycle by increasing flux through the glyoxylate shunt, and requires enzymes of the anaplerotic node and methylcitrate cycle. Targeting oxidative phosphorylation (OXPHOS) with BDQ and simultaneously inhibiting substrate level phosphorylation via genetic disruption of glycolysis leads to rapid sterilization. Our findings provide insight into the metabolic mechanism of BDQ-induced cell death and establish a paradigm for the development of combination therapies that target OXPHOS and glycolysis.
    Borah K, Rickman OJ, Voutsina N, Baple EL, Dias IH, Crosby AH, Griffiths HR (2020). Datasets of whole cell and mitochondrial oxysterols derived from THP-1, SH-SY5Y and human peripheral blood mononuclear cells using targeted metabolomics
    View abstract View full publication
    The raw datasets of oxysterol quantifications from whole cell and mitochondrial fractions of THP-1 monocytes and macrophages, neuronal-like SH-SH5Y cells and human peripheral blood mononuclear cells are presented. Oxysterols were quantified using a new liquid chromatography-mass spectrometry (LC-MS) and multiple reaction monitoring analysis published in the article "A quantitative LC-MS/MS method for analysis of mitochondrial-specific oxysterol metabolism" in Redox Biology [1]. This method showed improved extraction efficiency and recovery of mono and dihydroxycholesterols from cellular matrix. The datasets derived from the three cell lines are included in the appendix. These datasets provide new information about the oxysterol distribution in THP-1 monocytes and macrophages, SH-SY5Y cells and peripheral blood mononuclear cells. These datasets can be used as a guide for oxysterol distribution in the three cell lines for future studies, and can used for future method optimization, and for comparison of oxysterol recovery with other analytical techniques.
    Carolin C. M. Schulte, Khushboo Borah, Rachel M. Wheatley, Jason J. Terpolilli, Gerhard Saalbach, Nick Crang, Daan H. de Groot, R. George Ratcliffe, Nicholas J. Kruger, Antonis Papachristodoulou, Philip S. Poole (2021). Metabolic constraints on nitrogen fixation by rhizobia in legume nodules
    View abstract View full publication
    Rhizobia induce nodule formation on legume roots and differentiate into bacteroids, which use plant-derived dicarboxylates as energy and electron sources for reduction of atmospheric N2 into ammonia for secretion to plants. Using heterogeneous genome-scale datasets, we reconstructed a model of bacteroid metabolism to investigate the effects of varying dicarboxylate and oxygen supply on carbon and nitrogen allocation. Modelling and 13C metabolic flux analysis in bacteroids indicate that microaerobiosis restricts the decarboxylating arm of the TCA cycle and limits ammonia assimilation into glutamate. Catabolism of dicarboxylates induces a higher oxygen demand but also a higher NADH/NAD+ ratio compared to sugars. Carbon polymer synthesis and alanine secretion by bacteroids facilitate redox balance in microaerobic nodules with alanine secretion increasing as oxygen tension decreases. Our results provide a framework for understanding fundamental constraints on rhizobial metabolism during symbiotic nitrogen fixation.
    Khushboo Borah, Tom A. Mendum, Nathaniel D. Hawkins, Jane L. Ward, Michael H. Beale, Gerald Larrouy-Maumus, Apoorva Bhatt, Martine Moulin, Michael Haertlein, Gernot Strohmeier, Harald Pichler, V. Trevor Forsyth, View ORCID ProfileStephen Noack, Celia W. (2021). Metabolic flux partitioning between the TCA cycle and glyoxylate shunt combined with a reversible methyl citrate cycle provide nutritional flexibility for Mycobacterium tuberculosis
    View abstract View full publication
    The utilisation of multiple host-derived carbon substrates is required by  (Mtb) to successfully sustain a tuberculosis infection thereby identifying the Mtb specific metabolic pathways and enzymes required for carbon co-metabolism as potential drug targets. Metabolic flux represents the final integrative outcome of many different levels of cellular regulation that contribute to the flow of metabolites through the metabolic network. It is therefore critical that we have an in-depth understanding of the rewiring of metabolic fluxes in different conditions. Here, we employed 13C-metabolic flux analysis using stable isotope tracers (13C and 2H) and lipid fingerprinting to investigate the metabolic network of Mtb growing slowly on physiologically relevant carbon sources in a steady state chemostat. We demonstrate that Mtb is able to efficiently co-metabolise combinations of either cholesterol or glycerol along with C2 generating carbon substrates. The uniform assimilation of the carbon sources by Mtb throughout the network indicated no compartmentalization of metabolism in these conditions however there were substrate specific differences in metabolic fluxes. This work identified that partitioning of flux between the TCA cycle and the glyoxylate shunt combined with a reversible methyl citrate cycle as the critical metabolic nodes which underlie the nutritional flexibility of Mtb. These findings provide new insights into the metabolic architecture that affords adaptability of Mtb to divergent carbon substrates.
    Mycobacterium tuberculosis
    Khushboo Borah , Helen Griffiths (2020). Oxysterol_LCMS/MS raw data_whole cell/mitochondria
    Khushboo Borah, George Ratcliffe, Nicolas J Kruger, Philip s Poole (2016). Metabolic flux analysis of nitrogen-fixing bacteria
    K Borah, T Mendum, ND Hawkins, J Ward, M Beale, G Larrouy‐Maumus, A Bhatt, M Moulin, M Haertlein, G Strohmeier, H Pichler, T Forsyth, S Noack, C Goulding, J McFdden, D Beste (2021). Metabolic fluxes for nutritional flexibility of Mycobacterium tuberculosis
    View abstract View full publication
    The co‐catabolism of multiple host‐derived carbon substrates is required by  (Mtb) to successfully sustain a tuberculosis infection. However, the metabolic plasticity of this pathogen and the complexity of the metabolic networks present a major obstacle in identifying those nodes most amenable to therapeutic interventions. It is therefore critical that we define the metabolic phenotypes of Mtb in different conditions. We applied metabolic flux analysis using stable isotopes and lipid fingerprinting to investigate the metabolic network of Mtb growing slowly in our steady‐state chemostat system. We demonstrate that Mtb efficiently co‐metabolises either cholesterol or glycerol, in combination with two‐carbon generating substrates without any compartmentalisation of metabolism. We discovered that partitioning of flux between the TCA cycle and the glyoxylate shunt combined with a reversible methyl citrate cycle is the critical metabolic nodes which underlie the nutritional flexibility of Mtb. These findings provide novel insights into the metabolic architecture that affords adaptability of bacteria to divergent carbon substrates and expand our fundamental knowledge about the methyl citrate cycle and the glyoxylate shunt.
    Mycobacterium tuberculosis
    Carolin C. M. Schulte†, Khushboo Borah, Rachel M. Wheatly, Jason J. Terpolilli, Gerald Salbach, Nick Crang, Dan de Groot, R. George Ratcliffe,Nicholas J. Kruger, Antonis Papachristodoulou, Philip S Poole (2021). Metabolic control of nitrogen fixation in rhizobium-legume symbioses
    View abstract View full publication
    Rhizobia induce nodule formation on legume roots and differentiate into bacteroids, which catabolize plant-derived dicarboxylates to reduce atmospheric N2 into ammonia. Despite the agricultural importance of this symbiosis, the mechanisms that govern carbon and nitrogen allocation in bacteroids and promote ammonia secretion to the plant are largely unknown. Using a metabolic model derived from genome-scale datasets, we show that carbon polymer synthesis and alanine secretion by bacteroids facilitate redox balance in microaerobic nodules. Catabolism of dicarboxylates induces not only a higher oxygen demand but also a higher NADH/NAD+ ratio than sugars. Modeling and 13C metabolic flux analysis indicate that oxygen limitation restricts the decarboxylating arm of the tricarboxylic acid cycle, which limits ammonia assimilation into glutamate. By tightly controlling oxygen supply and providing dicarboxylates as the energy and electron source donors for N2 fixation, legumes promote ammonia secretion by bacteroids. This is a defining feature of rhizobium-legume symbioses.