title: Reexamination of the Physiological Role of PykA in Escherichia coli Revealed that It Negatively Regulates the Intracellular ATP Levels under Anaerobic Conditions.
abstract: Pyruvate kinase is one of the three rate-limiting glycolytic enzymes that catalyze the last step of glycolysis, conversion of phosphoenolpyruvate (PEP) into pyruvate, which is associated with ATP generation. Two isozymes of pyruvate kinase, PykF and PykA, are identified inEscherichia coliPykF is considered important, whereas PykA has a less-defined role. Prior studies inactivated thepykAgene to increase the level of its substrate, PEP, and thereby increased the yield of end products derived from PEP. We were surprised when we found apykA::Tn5mutant in a screen for increased yield of an end product derived from pyruvate (n-butanol), suggesting that the role of PykA needs to be reexamined. We show that thepykAmutant exhibited elevated intracellular ATP levels, biomass concentrations, glucose consumption, andn-butanol production. We also discovered that thepykAmutant expresses higher levels of a presumed pyruvate transporter, YhjX, permitting the mutant to recapture and metabolize excreted pyruvate. Furthermore, we demonstrated that the nucleotide diphosphate kinase activity of PykA leads to negative regulation of the intracellular ATP levels. Taking the data together, we propose that inactivation ofpykAcan be considered a general strategy to enhance the production of pyruvate-derived metabolites under anaerobic conditions.IMPORTANCEThis study showed that knocking outpykAsignificantly increased the intracellular ATP level and thus significantly increased the levels of glucose consumption, biomass formation, and pyruvate-derived product formation under anaerobic conditions.pykAwas considered to be encoding a dispensable pyruvate kinase; here we show thatpykAnegatively regulates the anaerobic glycolysis rate through regulating the energy distribution. Thus, knocking outpykAcan be used as a general strategy to increase the level of pyruvate-derived fermentative products.
authors.affiliation: CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China.|University of Chinese Academy of Sciences, Beijing, China.
authors.forename: Chunhua
authors.initials: C
authors.lastname: Zhao
authors.affiliation: CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China.|University of Chinese Academy of Sciences, Beijing, China.
authors.forename: Zhao
authors.initials: Z
authors.lastname: Lin
authors.affiliation: CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China redarmy305@gmail.com yli@im.ac.cn.
authors.forename: Hongjun
authors.initials: H
authors.lastname: Dong
authors.affiliation: CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China.
authors.forename: Yanping
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China redarmy305@gmail.com yli@im.ac.cn.
authors.forename: Yin
authors.initials: Y
authors.lastname: Li
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000693:Anaerobiosis; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D015971:Gene Expression Regulation, Enzymologic; D005947:Glucose / Q000378:metabolism; D010728:Phosphoenolpyruvate / Q000378:metabolism; D011770:Pyruvate Kinase / Q000235:genetics / Q000378:metabolism*
chemical_list: D029968:Escherichia coli Proteins; D010728:Phosphoenolpyruvate; D000255:Adenosine Triphosphate; D011770:Pyruvate Kinase; D005947:Glucose
keywords: ATP; Escherichia coli; PykA; anaerobic condition; physiological role