Archives of Microbiology

, Volume 152, Issue 6, pp 584–588

Isolation and characterization of oxaloacetate decarboxylase of Salmonella typhimurium, a sodium ion pump

  • Klaus Wifling
  • Peter Dimroth
Original Papers

Abstract

Anaerobic growth of Salmonella typhimurium on citrate is Na+-dependent and requires induction of the necessary enzymes during a 20–40 h lag phase. The citrate fermentation pathway involves citrate lyase and oxaloacetate decarboxylase. The decarboxylase is a membrane-bound. Na+-activated, biotin-containing enzyme that functions as a Na+ pump. Oxaloacetate decarboxylase was isolated by affinity chromatography of a Triton X-100 extract of the bacterial membranes on avidin-Sepharose. The enzyme consists of three subunits α, β, γ, with apparent molecular weights of 63800, 34500 and 10600. The α-chain contains a covalently attached biotin group and binds to antibodies raised against the α-subunit of oxaloacetate decarboxylase from Klebsiella pneumoniae. The Na+ transport function was reconstituted by incorporation of the puriried enzyme into proteoliposomes.

Key words

Citrate fermentation Oxaloacetate decarboxylase Salmonella typhimurium Sodium transport 

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References

  1. Amarasingham CR, Davis BD (1965) Regulation of α-ketoglutarate dehydrogenase formation in Escherichia coli. J Biol Chem 240:3664–3668Google Scholar
  2. Dagley S (1954) Dissimilation of citric acid by Aerobacter aerogenes and Escherichia coli. J Gen Microbiol 11:218–227Google Scholar
  3. Dimroth P (1982a) The generation of an electrochemical gradient of sodium ions upon decarboxylation of oxaloacetate by the membrane-bound and Na+-activated oxaloacetate decarboxylase from Klebsiella aerogenes. Eur J Biochem 121:443–449Google Scholar
  4. Dimroth P (1982b) The role of biotin and sodium in the decarboxylation of oxaloacetate by the membrane-bound oxaloacetate decarboxylase from Klebsiella aerogenes. Eur J Biochem 121:435–441Google Scholar
  5. Dimroth P (1986) Preparation, characterization and reconstitution of oxaloacetate decarboxylase from Klebsiella aerogenes, a sodium pump. Meth Enzymol 125:530–540Google Scholar
  6. Dimroth P (1987) Sodium ion transport decarboxylases and other aspects of sodium ion cycling in bacteria. Microbiol Rev 51:320–340Google Scholar
  7. Dimroth P, Thomer A (1983) Subunit composition of oxaloacetate decarboxylase and characterization of the α-chain as carboxyltransferase. Eur J Biochem 137:107–112Google Scholar
  8. Dimroth P, Thomer A (1986) Citrate transport in Klebsiella pneumoniae. Biol Chem Hoppe-Seyler 367:813–823Google Scholar
  9. Dimroth P, Thomer A (1988) Dissociation of the sodium-ion translocating oxaloacetate decarboxylase of Klebsiella pneumoniae and reconstitution of the active complex from the isolated subunits. Eur J Biochem 175:175–180Google Scholar
  10. Dimroth P, Thomer A (1989) A primary respiratory Na+ pump of an anaerobic bacterium: The Na+-dependent NADH: quinone oxidoreductase of Klebsiella pneumoniae. Arch Microbiol 151:439–444Google Scholar
  11. Hilpert W, Dimroth P (1983) Purification and characterization of a new sodium-transport decarboxylase. Methylmalonyl-CoA decarboxylase from Veillonella alcalescens. Eur J Biochem 132:579–587Google Scholar
  12. Kay W, Cameron M (1978) Citrate transport in Salmonella typhimurium. Arch Biochem Biophys 190:270–280Google Scholar
  13. Kröger A, Schimkat M, Niedermaier S (1974) Electrontransport phosphorylation coupled to fumarate reduction in anaerobically grown Proteus rettgeri. Biochim Biophys Acta 347:273–289Google Scholar
  14. Laemmli UK (1970) Clevage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685Google Scholar
  15. Laußermair E, Schwarz E, Oesterhelt D, Reinke H, Beyreuther K, Dimroth P (1989) The sodium ion translocating oxaloacetate decarboxylase of Klebsiella pneumoniae. Sequence of the integral membrane-bound subunits β and γ. J Biol Chem (in press)Google Scholar
  16. Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193:265–275Google Scholar
  17. Lütgens M, Gottschalk G (1980) Why a co-substrate is required for anaerobic growth of Escherichia coli on citrate. J Gen Microbiol 119:63–70Google Scholar
  18. O'Brien RW, Frost GM, Stern JR (1969) Enzymatic analysis of the requirement for sodium in aerobic growth of Salmonella typhimurium on citrate. J Bacteriol 99: 395–400Google Scholar
  19. Schwarz E, Oesterhelt D, Reinke H, Beyreuther K, Dimroth P (1988) The sodium ion translocating oxaloacetate decarboxylase of Klebsiella pneumoniae. Sequence of the biotin-containing α-subunit and relationship to other biotin-containing enzymes. J Biol Chem 263:9640–9645Google Scholar
  20. Stern JR (1967) Oxaloacetate decarboxylase of Aerobacter aerogenes. Inhibition by avidin and requirement for sodium ion. Biochemistry 6:3545–3551Google Scholar
  21. Towbin H, Staehelin T, GordonJ (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: Procedure and some applications. Proc Natl Acad Sci USA 76:4350–4354Google Scholar
  22. Tsang VCW, Peralta JM, Simons AR (1983) Enzyme-linked immunoelectrotransfer blot techniques (EITB) for studying the specificities of antigens and antibodies separated by gel electrophoresis. Meth Enzymol 92:377–391Google Scholar

Copyright information

© Springer-Verlag 1989

Authors and Affiliations

  • Klaus Wifling
    • 1
  • Peter Dimroth
    • 1
  1. 1.Institut für Physiologische Chemie der Technischen Universität MünchenMünchen 40Germany

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