Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
권호 표지
DOI QR코드

DOI QR Code

The ybcF Gene of Escherichia coli Encodes a Local Orphan Enzyme, Catabolic Carbamate Kinase

  • Nam Yeun Kim (Department of Life Science, Division of EcoScience, Ewha Womans University) ;
  • Ok Bin Kim (Department of Life Science, Division of EcoScience, Ewha Womans University)
  • Received : 2022.10.21
  • Accepted : 2022.11.01
  • Published : 2022.12.28
Download PDF
⟨ Previous Next ⟩

Abstract

Escherichia coli can use allantoin as its sole nitrogen source under anaerobic conditions. The ureidoglycolate produced by double release of ammonia from allantoin can flow into either the glyoxylate shunt or further catabolic transcarbamoylation. Although the former pathway is well studied, the genes of the latter (catabolic) pathway are not known. In the catabolic pathway, ureidoglycolate is finally converted to carbamoyl phosphate (CP) and oxamate, and then CP is dephosphorylated to carbamate by a catabolic carbamate kinase (CK), whereby ATP is formed. We identified the ybcF gene in a gene cluster containing fdrA-ylbE-ylbF-ybcF that is located downstream of the allDCE-operon. Reverse transcription PCR of total mRNA confirmed that the genes fdrA, ylbE, ylbF, and ybcF are co-transcribed. Deletion of ybcF caused only a slight increase in metabolic flow into the glyoxylate pathway, probably because CP was used to de novo synthesize pyrimidine and arginine. The activity of the catabolic CK was analyzed using purified YbcF protein. The Vmax is 1.82 U/mg YbcF for CP and 1.94 U/mg YbcF for ADP, and the KM value is 0.47 mM for CP and 0.43 mM for ADP. With these results, it was experimentally revealed that the ybcF gene of E. coli encodes catabolic CK, which completes anaerobic allantoin degradation through substrate-level phosphorylation. Therefore, we suggest renaming the ybcF gene as allK.

Keywords

Acknowledgement

This study was funded by the Basic Science Research Program through the National Research Foundation of Korea (NRF-2021R1A6A1A10039823 and NRF-2019R1A2C1008066), and was supported by RP-Grant 2020 of Ewha Womans University.

References

  1. Jones ME, Lipmann F. 1960. Chemical and enzymatic synthesis of carbamyl phosphate. Proc. Natl. Acad. Sci. USA 4: 1194-1205. https://doi.org/10.1073/pnas.46.9.1194
  2. Jones ME. 1963. Carbamyl phosphate: many forms of life use this molecule to synthesize arginine, uracil, and adenosine triphosphate. Science 140: 1373-1379. https://doi.org/10.1126/science.140.3574.1373
  3. Shi D, Caldovic L, Tuchman M. 2018. Sources and fates of carbamyl phosphate: a labile energy-rich molecule with multiple facets. Biology 7: 34.
  4. Walsh CT, Tu BP, Tang Y. 2018. Eight kinetically stable but thermodynamically activated molecules that power cell metabolism. Chem. Rev. 118: 1460-1494. https://doi.org/10.1021/acs.chemrev.7b00510
  5. Trotta PP, Burt ME, Haschemeyer RH, Meister A. 1971. Reversible dissociation of carbamyl phosphate synthetase into a regulated synthesis subunit and a subunit required for glutamine utilization. Proc. Natl. Acad. Sci. USA 68: 2599-2603. https://doi.org/10.1073/pnas.68.10.2599
  6. Purcarea C, Simon V, Prieur D, Herve G. 1996. Purification and characterization of carbamoyl-phosphate synthetase from the deep-sea hyperthermophilic archaebacterium Pyrococcus abyssi. Eur. J. Biochem. 236: 189-199. https://doi.org/10.1111/j.1432-1033.1996.00189.x
  7. Durbecq V, Legrain C, Roovers M, Pierard A, Glansdorff N. 1997. The carbamate kinase-like carbamoyl phosphate synthetase of the hyperthermophilic archaeon Pyrococcus furiosus, a missing link in the evolution of carbamoyl phosphate biosynthesis. Proc. Natl. Acad. Sci. USA 94: 12803-12808. https://doi.org/10.1073/pnas.94.24.12803
  8. Ramon-Maiques S, Marina A, Uriarte M, Fita I, Rubio V. 2000. The 1.5 A resolution crystal structure of the carbamate kinase-like carbamoyl phosphate synthetase from the hyperthermophilic archaeon Pyrococcus furiosus, bound to ADP, confirms that this thermostable enzyme is a carbamate kinase, and provides insight into substrate binding and stability in carbamate kinases. J. Mol. Biol. 299: 463-476. https://doi.org/10.1006/jmbi.2000.3779
  9. Luthi E, Mercenier A, Haas D. 1986. The arcABC operon required for fermentative growth of Pseudomonas aeruginosa on arginine: Tn5-751-assisted cloning and localization of structural genes. J. Gen. Microbiol. 132: 2667-2675.
  10. Barcelona-Andres B, Marina A, Rubio V. 2002. Gene structure, organization, expression, and potential regulatory mechanisms of arginine catabolism in Enterococcus faecalis. J Bacteriol. 184: 6289-6300. https://doi.org/10.1128/JB.184.22.6289-6300.2002
  11. Noens EE, Lolkema JS. 2017. Convergent evolution of the arginine deiminase pathway: the ArcD and ArcE arginine/ornithine exchangers. Microbiologyopen 6: e00412.
  12. Majsnerowska M, Noens EEE, Lolkema JS. 2018. Arginine and citrulline catabolic pathways encoded by the arc gene cluster of Lactobacillus brevis ATCC 367. J. Bacteriol. 200: e00182-18.
  13. Simon JP, Stalon V. 1982. Enzymes of agmatine degradation and the control of their synthesis in Streptococcus faecalis. J. Bacteriol. 152: 676-681. https://doi.org/10.1128/jb.152.2.676-681.1982
  14. Griswold AR, Jameson-Lee M, Burne RA. 2006. Regulation and physiologic significance of the agmatine deiminase system of Streptococcus mutans UA159. J. Bacteriol. 188: 834-841. https://doi.org/10.1128/JB.188.3.834-841.2006
  15. Li Y, Jin Z, Yu X, Allewell NM, Tuchman M, Shi D. 2011. The ygeW encoded protein from Escherichia coli is a knotted ancestral catabolic transcarbamylase. Proteins 79: 2327-2334. https://doi.org/10.1002/prot.23043
  16. Kim NY, Lee YJ, Park JW, Kim SN, Kim EY, Kim Y, et al. 2021. An Escherichia coli FdrA variant derived from syntrophic coculture with a methanogen increases succinate production due to changes in allantoin degradation. mSphere 6: e0065421.
  17. Smith AA, Belda E, Viari A, Medigue C, Vallenet D. 2012. The CanOE strategy: integrating genomic and metabolic contexts across multiple prokaryote genomes to find candidate genes for orphan enzymes. PLoS Comput. Biol. 8: e1002540.
  18. Miller JH. 1972. Experiments in Molecular Genetics. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York.
  19. Neidhardt FC, Bloch PL, Smith DF. 1974. Culture medium for enterobacteria. J. Bacteriol. 119: 736-747. https://doi.org/10.1128/jb.119.3.736-747.1974
  20. Datsenko KA, Wanner BL. 2000. One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc. Natl. Acad. Sci. USA 97: 6640-6645. https://doi.org/10.1073/pnas.120163297
  21. Hao Lin, En-Ze Deng, Hui Ding, Wei Chen, Kuo-Chen Chou. 2014. iPro54-PseKNC: a sequence-based predictor for identifying sigma-54 promoters in prokaryote with pseudo k-tuple nucleotide composition. Nucleic Acids Res. 42: 12961-12972. https://doi.org/10.1093/nar/gku1019
  22. Rahman MS, Aktar U, Jani MR, Shatabda S. 2019. iPro70-FMWin: identifying Sigma70 promoters using multiple windowing and minimal features. Mol Genet. Genomics 294: 69-84. https://doi.org/10.1007/s00438-018-1487-5
  23. Valentine RC, Bojanowski R, Gaudy E, Wolfe RS. 1962. Mechanism of the allantoin fermentation. J. Biol. Chem. 237: 2271-2277. https://doi.org/10.1016/S0021-9258(19)63431-9
  24. Chen CZ, Southall N, Galkin A, Lim K, Marugan JJ, Kulakova L, et al. 2012. A homogenous luminescence assay reveals novel inhibitors for giardia lamblia carbamate kinase. Curr. Chem. Genomics 6: 93-102. https://doi.org/10.2174/1875397301206010093
  25. Abdelal AT. 1979. Arginine catabolism by microorganisms. Annu. Rev. Microbiol. 33: 139-168. https://doi.org/10.1146/annurev.mi.33.100179.001035
  26. Hering S, Sieg A, Kreikemeyer B, Fiedler T. 2013. Kinetic characterization of arginine deiminase and carbamate kinase from Streptococcus pyogenes M49. Protein Expr. Purif. 91: 61-68. https://doi.org/10.1016/j.pep.2013.07.002
  27. Marina A, Uriarte M, Barcelona B, Fresquet V, Cervera J, Rubio V. 1998. Carbamate kinase from Enterococcus faecalis and Enterococcus faecium--cloning of the genes, studies on the enzyme expressed in Escherichia coli, and sequence similarity with N-acetyl-L-glutamate kinase. Eur. J. Biochem. 253: 280-291. https://doi.org/10.1046/j.1432-1327.1998.2530280.x
  28. Abdelal AT, Bibb WF, Nainan O. 1982. Carbamate kinase from Pseudomonas aeruginosa: purification, characterization, physiological role, and regulation. J. Bacteriol. 151: 1411-1419. https://doi.org/10.1128/jb.151.3.1411-1419.1982
  29. Manca de Nadra MC, Nadra Chaud CA, Pesce de Ruiz Holgado A, Oliver G. 1986. Carbamate kinase of Lactobacillus buchneri NCDO110. I. Purification and properties. Biotechnol. Appl. Biochem. 8: 46-52.
  30. Galkin A, Kulakova L, Wu R, Nash TE, Dunaway-Mariano D, Herzberg O. 2010. X-ray structure and characterization of carbamate kinase from the human parasite Giardia lamblia. Acta Crystallogr. Sect. F Struct. Biol. Cryst. Commun. 66: 386-390. https://doi.org/10.1107/S1744309110004665
  31. Minotto L, Tutticci EA, Bagnara AS, Schofield PJ, Edwards MR. 1999. Characterisation and expression of the carbamate kinase gene from Giardia intestinalis. Mol. Biochem. Parasitol. 98: 43-51. https://doi.org/10.1016/S0166-6851(98)00141-8
  32. Minotto L, Edwards MR, Bagnara AS. 2000. Trichomonas vaginalis: characterization, expression, and phylogenetic analysis of a carbamate kinase gene sequence. Exp. Parasitol. 95: 54-62. https://doi.org/10.1006/expr.2000.4507
  33. Cusa E, Obradors N, Baldoma L, Badia J, Aguilar J. 1999. Genetic analysis of a chromosomal region containing genes required for assimilation of allantoin nitrogen and linked glyoxylate metabolism in Escherichia coli. J. Bacteriol. 181: 7479-7484. https://doi.org/10.1128/JB.181.24.7479-7484.1999
  34. Manca de Nadra MC, Pesce de Ruiz Holgado AA, Oliver G. 1987. Carbamate kinase of Lactobacillus buchneri NCDO110. II. Kinetic studies and reaction mechanism. Biotechnol. Appl. Biochem. 9: 141-145.
  35. Stebbins JW, Xu W, Kantrowitz ER. 1989. Three residues involved in binding and catalysis in the carbamyl phosphate binding site of Escherichia coli aspartate transcarbamylase. Biochemistry 28: 2592-600. https://doi.org/10.1021/bi00432a037
  36. Kuo LC, Lipscomb WN, Kantrowitz ER. 1982. Zn(II)-induced cooperativity of Escherichia coli ornithine transcarbamoylase. Proc. Natl. Acad. Sci. USA 79: 2250-2254. https://doi.org/10.1073/pnas.79.7.2250
  37. Baur H, Tricot C, Stalon V, Haas D. 1990. Converting catabolic ornithine carbamoyltransferase to an anabolic enzyme. J. Biol. Chem. 265: 14728-14731. https://doi.org/10.1016/S0021-9258(18)77171-8