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Polyphosphate/ATP-dependent NAD kinase of Corynebacterium glutamicum: biochemical properties and impact of ppnK overexpression on lysine production

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Abstract

Nicotinamide adenine dinucleotide phosphate (NADP) is synthesized by phosphorylation of either oxidized or reduced nicotinamide adenine dinucleotide (NAD/NADH). Here, the cg1601/ppnK gene product from Corynebacterium glutamicum genome was purified from recombinant Escherichia coli and enzymatic characterization revealed its activity as a polyphosphate (PolyP)/ATP-dependent NAD kinase (PPNK). PPNK from C. glutamicum was shown to be active as homotetramer accepting PolyP, ATP, and even ADP for phosphorylation of NAD. The catalytic efficiency with ATP as phosphate donor for phosphorylation of NAD was higher than with PolyP. With respect to the chain length of PolyP, PPNK was active with short-chain PolyPs. PPNK activity was independent of bivalent cations when using ATP, but was enhanced by manganese and in particular by magnesium ions. When using PolyP, PPNK required bivalent cations, preferably manganese ions, for activity. PPNK was inhibited by NADP and NADH at concentrations below millimolar. Overexpression of ppnK in C. glutamicum wild type slightly reduced growth and ppnK overexpression in the lysine producing strain DM1729 resulted in a lysine product yield on glucose of 0.136 ± 0.006 mol lysine (mol glucose)−1, which was 12% higher than that of the empty vector control strain.

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References

  • Abe S, Takayarna K, Kinoshita S (1967) Taxonomical studies on glutamic acid producing bacteria. J Gen Appl Microbiol 13:279–301

    Article  Google Scholar 

  • Ando S, Ochiai K, Yokoi H, Hashimoto S, Yonetani Y (2002) Novel glucose-6-phosphate dehydrogenase. Patent WO0198472 (2002-01-02)

  • Antelmann H, Scharf C, Hecker M (2000) Phosphate starvation-inducible proteins of Bacillus subtilis: proteomics and transcriptional analysis. J Bacteriol 182:4478–4490

    Article  CAS  Google Scholar 

  • Bakali HM, Herman MD, Johnson KA, Kelly AA, Wieslander A, Hallberg BM, Nordlund P (2007) Crystal structure of YegS, a homologue to the mammalian diacylglycerol kinases, reveals a novel regulatory metal binding site. J Biol Chem 282:19644–19652

    Article  CAS  Google Scholar 

  • Becker J, Klopprogge C, Herold A, Zelder O, Bolten CJ, Wittmann C (2007) Metabolic flux engineering of l-lysine production in Corynebacterium glutamicum—over expression and modification of G6P dehydrogenase. J Biotechnol 132:99–109

    Article  CAS  Google Scholar 

  • Berrin JG, Pierrugues O, Brutesco C, Alonso B, Montillet JL, Roby D, Kazmaier M (2005) Stress induces the expression of AtNADK-1, a gene encoding a NAD(H) kinase in Arabidopsis thaliana. Mol Genet Genomics 273:10–19

    Article  CAS  Google Scholar 

  • Börmann ER, Eikmanns BJ, Sahm H (1992) Molecular analysis of the Corynebacterium glutamicum gdh gene-encoding glutamate dehydrogenase. Mol Microbiol 6:317–326

    Article  Google Scholar 

  • Butler JR, McGuinness ET (1982) Candida utilis NAD+ kinase: purification, properties and affinity gel studies. Int J Biochem 14:839–844

    Article  CAS  Google Scholar 

  • Chai MF, Chen QJ, An R, Chen YM, Chen J, Wang XC (2005) NADK2, an Arabidopsis chloroplastic NAD kinase, plays a vital role in both chlorophyll synthesis and chloroplast protection. Plant Mol Biol 59:553–564

    Article  CAS  Google Scholar 

  • Cremer J, Treptow C, Eggeling L, Sahm H (1988) Regulation of enzymes of lysine biosynthesis in Corynebacterium glutamicum. J Gen Microbiol 134:3221–3229

    CAS  Google Scholar 

  • Docampo R (2006) Acidocalcisomes and polyphosphate granules. In: Shively JM (ed) Inclusions in prokaryotes vol 1. Springer, Berlin Germany, pp 53–70

    Chapter  Google Scholar 

  • Dominguez H, Rollin C, Guyonvarch A, Guerquin-Kern JL, Cocaign-Bousquet M, Lindley ND (1998) Carbon-flux distribution in the central metabolic pathways of Corynebacterium glutamicum during growth on fructose. Eur J Biochem 254:96–102

    Article  CAS  Google Scholar 

  • Eggeling L, Bott M (eds) (2005) Handbook of Corynebacterium glutamicum. CRC Press LLC, Boca Raton

    Google Scholar 

  • Eikmanns BJ, Rittmann D, Sahm H (1995) Cloning, sequence analysis, expression, and inactivation of the Corynebacterium glutamicum icd gene encoding isocitrate dehydrogenase and biochemical characterization of the enzyme. J Bacteriol 177:774–782

    CAS  Google Scholar 

  • Garavaglia S, Galizzi A, Rizzi M (2003) Allosteric regulation of Bacillus subtilis NAD kinase by quinolinic acid. J Bacteriol 185:4844–4850

    Article  CAS  Google Scholar 

  • Georgi T, Rittmann D, Wendisch VF (2005) Lysine and glutamate production by Corynebacterium glutamicum on glucose, fructose and sucrose: roles of malic enzyme and fructose-1, 6-bisphosphatase. Metab Eng 7:291–301

    Article  CAS  Google Scholar 

  • Gourdon P, Baucher MF, Lindley ND, Guyonvarch A (2000) Cloning of the malic enzyme gene from Corynebacterium glutamicum and role of the enzyme in lactate metabolism. Appl Environ Microbiol 66:2981–2987

    Article  CAS  Google Scholar 

  • Hanahan D (1985) Techniques for transformation of E. coli. In: Glover DM (ed) DNA cloning: a practical approach, vol 1. IRL Press, Oxford, pp 109–135

    Google Scholar 

  • Hoischen C, Kramer R (1989) Evidence for an efflux carrier system involved in the secretion of glutamate by Corynebacterium glutamicum. Arch Microbiol 151:342–347

    Article  CAS  Google Scholar 

  • Ishige T, Krause M, Bott M, Wendisch VF, Sahm H (2003) The phosphate starvation stimulon of Corynebacterium glutamicum determined by DNA microarray analyses. J Bacteriol 185:4519–4529

    Article  CAS  Google Scholar 

  • Kabus A, Georgi T, Wendisch VF, Bott M (2007) Expression of the Escherichia coli pntAB genes encoding a membrane-bound transhydrogenase in Corynebacterium glutamicum improves l-lysine formation. Appl Microbiol Biotechnol 75:47–53

    Article  CAS  Google Scholar 

  • Kalinowski J, Bathe B, Bartels D, Bischoff N, Bott M, Burkovski A, Dusch N, Eggeling L, Eikmanns BJ, Gaigalat L, Goesmann A, Hartmann M, Huthmacher K, Kramer R, Linke B, McHardy AC, Meyer F, Mockel B, Pfefferle W, Puhler A, Rey DA, Ruckert C, Rupp O, Sahm H, Wendisch VF, Wiegrabe I, Tauch A (2003) The complete Corynebacterium glutamicum ATCC 13032 genome sequence and its impact on the production of l-aspartate-derived amino acids and vitamins. J Biotechnol 104:5–25

    Article  CAS  Google Scholar 

  • Kawai S, Mori S, Mukai T, Suzuki S, Yamada T, Hashimoto W, Murata K (2000) Inorganic Polyphosphate/ATP-NAD kinase of Micrococcus flavus and Mycobacterium tuberculosis H37Rv. Biochem Biophys Res Commun 276:57–63

    Article  CAS  Google Scholar 

  • Kawai S, Mori S, Mukai T, Hashimoto W, Murata K (2001a) Molecular characterization of Escherichia coli NAD kinase. Eur J Biochem 268:4359–4365

    Article  CAS  Google Scholar 

  • Kawai S, Suzuki S, Mori S, Murata K (2001b) Molecular cloning and identification of UTR1 of a yeast Saccharomyces cerevisiae as a gene encoding an NAD kinase. FEMS Microbiol Lett 200:181–184

    Article  CAS  Google Scholar 

  • Klauth P, Pallerla SR, Vidaurre D, Ralfs C, Wendisch VF, Schoberth SM (2006) Determination of soluble and granular inorganic polyphosphate in Corynebacterium glutamicum. Appl Microbiol Biotechnol 72:1099–1106

    Article  CAS  Google Scholar 

  • Kobayashi K, Ehrlich SD, Albertini A, Amati G, Andersen KK, Arnaud M, Asai K, Ashikaga S, Aymerich S, Bessieres P, Boland F, Brignell SC, Bron S, Bunai K, Chapuis J, Christiansen LC, Danchin A, Debarbouille M, Dervyn E, Deuerling E, Devine K, Devine SK, Dreesen O, Errington J, Fillinger S, Foster SJ, Fujita Y, Galizzi A, Gardan R, Eschevins C, Fukushima T, Haga K, Harwood CR, Hecker M, Hosoya D, Hullo MF, Kakeshita H, Karamata D, Kasahara Y, Kawamura F, Koga K, Koski P, Kuwana R, Imamura D, Ishimaru M, Ishikawa S, Ishio I, Le Coq D, Masson A, Mauel C, Meima R, Mellado RP, Moir A, Moriya S, Nagakawa E, Nanamiya H, Nakai S, Nygaard P, Ogura M, Ohanan T, O’Reilly M, O’Rourke M, Pragai Z, Pooley HM, Rapoport G, Rawlins JP, Rivas LA, Rivolta C, Sadaie A, Sadaie Y, Sarvas M, Sato T, Saxild HH, Scanlan E, Schumann W, Seegers JF, Sekiguchi J, Sekowska A, Seror SJ, Simon M, Stragier P, Studer R, Takamatsu H, Tanaka T, Takeuchi M, Thomaides HB, Vagner V, van Dijl JM, Watabe K, Wipat A, Yamamoto H, Yamamoto M, Yamamoto Y, Yamane K, Yata K, Yoshida K, Yoshikawa H, Zuber U, Ogasawara N (2003) Essential Bacillus subtilis genes. Proc Natl Acad Sci USA 100:4678–4683

    Article  CAS  Google Scholar 

  • Kocan M, Schaffer S, Ishige T, Sorger-Herrmann U, Wendisch VF, Bott M (2006) Two-component systems of Corynebacterium glutamicum: deletion analysis and involvement of the PhoS-PhoR system in the phosphate starvation response. J Bacteriol 188:724–732

    Article  CAS  Google Scholar 

  • Kornberg A (1950) Enzymatic synthesis of triphosphopyridine nucleotide. J Biol Chem 182:805–813

    CAS  Google Scholar 

  • Kornberg A, Rao NN, Ault-Riche D (1999) Inorganic polyphosphate: a molecule of many functions. Annu Rev Biochem 68:89–125

    Article  CAS  Google Scholar 

  • Kulaev IS, Vagabov VM, Kulakovskaya TV (2004) The biochemistry of inorganic polyphosphates, 2nd edn. Wiley, Chichester

    Google Scholar 

  • Lerner F, Niere M, Ludwig A, Ziegler M (2001) Structural and functional characterization of human NAD kinase. Biochem Biophys Res Commun 288:69–74

    Article  CAS  Google Scholar 

  • Lindner SN, Vidaurre D, Willbold S, Schoberth SM, Wendisch VF (2007) NCgl2620 encodes a class II polyphosphate kinase in Corynebacterium glutamicum. Appl Environ Microbiol 73:5026–5033

    Article  CAS  Google Scholar 

  • Lindner SN, Knebel S, Wesseling H, Schoberth SM, Wendisch VF (2009) Exopolyphosphatases PPX1 and PPX2 from Corynebacterium glutamicum. Appl Environ Microbiol 75:3161–3170

    Article  CAS  Google Scholar 

  • Liu J, Lou Y, Yokota H, Adams PD, Kim R, Kim SH (2005) Crystal structures of an NAD kinase from Archaeoglobus fulgidus in complex with ATP, NAD, or NADP. J Mol Biol 354:289–303

    Article  CAS  Google Scholar 

  • Marx A, Hans S, Mockel B, Bathe B, de Graaf AA, McCormack AC, Stapleton C, Burke K, O’Donohue M, Dunican LK (2003) Metabolic phenotype of phosphoglucose isomerase mutants of Corynebacterium glutamicum. J Biotechnol 104:185–197

    Article  CAS  Google Scholar 

  • Mattson G, Conklin E, Desai S, Nielander G, Savage MD, Morgensen S (1993) A practical approach to crosslinking. Mol Biol Rep 17:167–183

    Article  CAS  Google Scholar 

  • McGuinness ET, Butler JR (1985) NAD+ kinase—a review. Int J Biochem 17:1–11

    Article  CAS  Google Scholar 

  • Mori S, Kawai S, Shi F, Mikami B, Murata K (2005a) Molecular conversion of NAD kinase to NADH kinase through single amino acid residue substitution. J Biol Chem 280:24104–24112

    Article  CAS  Google Scholar 

  • Mori S, Yamasaki M, Maruyama Y, Momma K, Kawai S, Hashimoto W, Mikami B, Murata K (2005b) NAD-binding mode and the significance of intersubunit contact revealed by the crystal structure of Mycobacterium tuberculosis NAD kinase-NAD complex. Biochem Biophys Res Commun 327:500–508

    Article  CAS  Google Scholar 

  • Moritz B, Striegel K, De Graaf AA, Sahm H (2000) Kinetic properties of the glucose-6-phosphate and 6-phosphogluconate dehydrogenases from Corynebacterium glutamicum and their application for predicting pentose phosphate pathway flux in vivo. Eur J Biochem 267:3442–3452

    Article  CAS  Google Scholar 

  • Moritz B, Striegel K, de Graaf AA, Sahm H (2002) Changes of pentose phosphate pathway flux in vivo in Corynebacterium glutamicum during leucine-limited batch cultivation as determined from intracellular metabolite concentration measurements. Metab Eng 4:295–305

    Article  CAS  Google Scholar 

  • Ochiai A, Mori S, Kawai S, Murata K (2004) Overexpression, purification, and characterization of ATP-NAD kinase of Sphingomonas sp. A1. Protein Expr Purif 36:124–130

    Article  CAS  Google Scholar 

  • Omumasaba CA, Okai N, Inui M, Yukawa H (2004) Corynebacterium glutamicum glyceraldehyde-3-phosphate dehydrogenase isoforms with opposite, ATP-dependent regulation. J Mol Microbiol Biotechnol 8:91–103

    Article  CAS  Google Scholar 

  • Outten CE, Culotta VC (2003) A novel NADH kinase is the mitochondrial source of NADPH in Saccharomyces cerevisiae. Embo J 22:2015–2024

    Article  CAS  Google Scholar 

  • Pallerla SR, Knebel S, Polen T, Klauth P, Hollender J, Wendisch VF, Schoberth SM (2005) Formation of volutin granules in Corynebacterium glutamicum. FEMS Microbiol Lett 243:133–140

    Article  CAS  Google Scholar 

  • Patek M, Bilic M, Krumbach K, Eikmanns B, Sahm H, Eggeling L (1997) Identification and transcriptional analysis of the dapB-ORF2-dapA-ORF4 operon of Corynebacterium glutamicum, encoding two enzymes involved in l-lysine synthesis. Biotechnol Lett 19:1113

    Article  CAS  Google Scholar 

  • Pollak N, Dolle C, Ziegler M (2007) The power to reduce: pyridine nucleotides—small molecules with a multitude of functions. Biochem J 402:205–218

    Article  CAS  Google Scholar 

  • Raffaelli N, Finaurini L, Mazzola F, Pucci L, Sorci L, Amici A, Magni G (2004) Characterization of Mycobacterium tuberculosis NAD kinase: functional analysis of the full-length enzyme by site-directed mutagenesis. Biochemistry 43:7610–7617

    Article  CAS  Google Scholar 

  • Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425

    CAS  Google Scholar 

  • Sakuraba H, Kawakami R, Ohshima T (2005) First archaeal inorganic polyphosphate/ATP-dependent NAD kinase, from hyperthermophilic archaeon Pyrococcus horikoshii: cloning, expression, and characterization. Appl Environ Microbiol 71:4352–4358

    Article  CAS  Google Scholar 

  • Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor

    Google Scholar 

  • Sassetti CM, Boyd DH, Rubin EJ (2003) Genes required for mycobacterial growth defined by high density mutagenesis. Mol Microbiol 48:77–84

    Article  CAS  Google Scholar 

  • Schaaf S, Bott M (2007) Target genes and DNA-binding sites of the response regulator PhoR from Corynebacterium glutamicum. J Bacteriol 189:5002–5011

    Article  CAS  Google Scholar 

  • Schröder HC, Müller WEG (1999) (ed) Inorganic polyphosphates: biochemistry, biology, biotechnology. Progress in molecular and subcellular biology, vol. 23. Springer, Berlin, Germany

  • Shi F, Kawai S, Mori S, Kono E, Murata K (2005) Identification of ATP-NADH kinase isozymes and their contribution to supply of NADP(H) in Saccharomyces cerevisiae. Febs J 272:3337–3349

    Article  CAS  Google Scholar 

  • Shimizu H, Hirasawa T (2007) Production of glutamate and glutamate-related amino acids: molecular mechanism analysis and metabolic engineering. In: Wendisch VF (ed) Amino acid biosynthesis—pathways, regulation and metabolic engineering. Springer, Heidelberg

    Google Scholar 

  • Stansen C, Uy D, Delaunay S, Eggeling L, Goergen JL, Wendisch VF (2005) Characterization of a Corynebacterium glutamicum lactate utilization operon induced during temperature-triggered glutamate production. Appl Environ Microbiol 71:5920–5928

    Article  CAS  Google Scholar 

  • Strand MK, Stuart GR, Longley MJ, Graziewicz MA, Dominick OC, Copeland WC (2003) POS5 gene of Saccharomyces cerevisiae encodes a mitochondrial NADH kinase required for stability of mitochondrial DNA. Eukaryot Cell 2:809–820

    Article  CAS  Google Scholar 

  • Studier FW, Moffatt BA (1986) Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes. J Mol Biol 189:113–130

    Article  CAS  Google Scholar 

  • Thompson JD, Higgins DG, Gibson TJ (1994) CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22:4673–4680

    Article  CAS  Google Scholar 

  • Turner WL, Waller JC, Vanderbeld B, Snedden WA (2004) Cloning and characterization of two NAD kinases from Arabidopsis. identification of a calmodulin binding isoform. Plant Physiol 135:1243–1255

    Article  CAS  Google Scholar 

  • Wellerdiek M, Winterhoff D, Reule W, Brandner J, Oldiges M (2009) Metabolic quenching of Corynebacterium glutamicum: efficiency of methods and impact of cold shock. Bioprocess Biosyst Eng 32:581–592

    Article  CAS  Google Scholar 

  • Wendisch VF, Bott M (2008) Phosphorus metabolism and its regulation. In: Burkovski A (ed) Corynebacteria: genomics and molecular biology. Caister Academic Press, Norfolk, pp 203–216

    Google Scholar 

  • Wittmann C, Becker J (2007) The l-lysine story: from metabolic pathways to industrial production. In: Wendisch VF (ed) Amino acid biosynthesis—pathways, regulation and metabolic engineering. Springer, Heidelberg

    Google Scholar 

  • Wittmann C, Kim HM, Heinzle E (2004) Metabolic network analysis of lysine producing Corynebacterium glutamicum at a miniaturized scale. Biotechnol Bioeng 87:1–6

    Article  CAS  Google Scholar 

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Authors and Affiliations

  1. Institute of Molecular Microbiology and Biotechnology, Westfalian Wilhelms University Muenster, 48149, Muenster, Germany

    Steffen N. Lindner, Henrike Niederholtmeyer & Katja Schmitz

  2. Institut für Biotechnologie 1, Forschungszentrum Jülich, 52425, Jülich, Germany

    Siegfried M. Schoberth

  3. Chair of Genetics Prokaryotes, Bielefeld University, Bielefeld, Universitätsstr. 25, 33615, Bielefeld, Germany

    Volker F. Wendisch

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  1. Steffen N. Lindner
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  2. Henrike Niederholtmeyer
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  3. Katja Schmitz
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  4. Siegfried M. Schoberth
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  5. Volker F. Wendisch
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Correspondence to Volker F. Wendisch.

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Steffen N. Lindner and Henrike Niederholtmeyer contributed equally to this paper.

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Lindner, S.N., Niederholtmeyer, H., Schmitz, K. et al. Polyphosphate/ATP-dependent NAD kinase of Corynebacterium glutamicum: biochemical properties and impact of ppnK overexpression on lysine production. Appl Microbiol Biotechnol 87, 583–593 (2010). https://doi.org/10.1007/s00253-010-2481-y

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