Abstract
Pyruvate kinase (PYK) is an essential glycolytic enzyme that controls glycolytic flux and is critical for ATP production in all organisms, with tight regulation by multiple metabolites. Yet the allosteric mechanisms governing PYK activity in bacterial pathogens are poorly understood. Here we report biochemical, structural and metabolomic evidence that Mycobacterium tuberculosis (Mtb) PYK uses AMP and glucose-6-phosphate (G6P) as synergistic allosteric activators that function as a molecular “OR logic gate” to tightly regulate energy and glucose metabolism. G6P was found to bind to a previously unknown site adjacent to the canonical site for AMP. Kinetic data and structural network analysis further show that AMP and G6P work synergistically as allosteric activators. Importantly, metabolome profiling in the Mtb surrogate, Mycobacterium bovis BCG, reveals significant changes in AMP and G6P levels during nutrient deprivation, which provides insights into how a PYK OR gate would function during the stress of Mtb infection.
Highlights
Mycobacterium tuberculosis (Mtb) is among the deadliest infectious diseases on a global scale, killing more than one-million people annually[1], with emerging antimicrobial drug resistance posing serious challenges to existing diagnosis and treatment programs[1]
pyruvate kinase (PYK) from higher organisms have a single essential modulator, with the exception of F16BP and amino-acid regulation of human M2PYK in cancer-cell proliferation[19,20,21]. This stands in contrast to the many bacterial PYKs that use ‘noncanonical’ effectors such as Adenosine monophosphate (AMP) and the sugar monophosphates glucose 6-phosphate (G6P) and ribose 5-phosphate (R5P) for allosteric regulation (Supplementary Table 1), including PYKs from important human pathogens such as Mtb[9], Streptococcus mutans[22], Staphylococcus aureus[23] and Salmonella typhimurium[16]
Allosteric mechanisms of key central carbon metabolism (CCM) enzymes enable bacteria to efficiently sense the changes in metabolite levels and react immediately by allosterically regulating mechanisms to maintain homoeostasis and defence against environmental threats[5]
Summary
Mycobacterium tuberculosis (Mtb) is among the deadliest infectious diseases on a global scale, killing more than one-million people annually[1], with emerging antimicrobial drug resistance posing serious challenges to existing diagnosis and treatment programs[1]. Increasing evidence suggests that the metabolic flexibility of central carbon metabolism (CCM: glycolysis, gluconeogenesis, pentose phosphate pathway and TCA pathway) is critical in Mtb physiology and pathogenicity[5,6,7] This is illustrated by the rapid regulation of glycolytic activity in response to changes in ATP levels, which explains the enhanced efficacy of drug combinations that target the electron transport chain[8]. PYKs from higher organisms have a single essential modulator, with the exception of F16BP and amino-acid regulation of human M2PYK in cancer-cell proliferation[19,20,21] This stands in contrast to the many bacterial PYKs that use ‘noncanonical’ effectors such as AMP and the sugar monophosphates glucose 6-phosphate (G6P) and ribose 5-phosphate (R5P) for allosteric regulation (Supplementary Table 1), including PYKs from important human pathogens such as Mtb[9], Streptococcus mutans[22], Staphylococcus aureus[23] and Salmonella typhimurium[16] (a sequence alignment of selected bacterial PYKs is shown in Supplementary Fig. 1 and pairwise identities in Supplementary Table 2). Stress-induced metabolomic changes in the Mtb surrogate, Mycobacterium bovis BCG, point to AMP and G6P as molecular input signals that position MtbPYK as a unique molecular OR ‘logic gate’ to sense changes in energy and sugar levels during Mtb infection
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
