Skip to main content
SpringerLink
Log in

Characterization of a Haloferax volcanii member of the enolase superfamily: deletion mutant construction, expression analysis, and transcriptome comparison

  • Original Paper
  • Published:
Archives of Microbiology Aims and scope Submit manuscript

Abstract

The enolase superfamily (COG4948) contains proteins with very different biological functions including regulators like the Escherichia coli RspA and metabolic enzymes like enolase. To unravel the biological function of an archaeal family member, an in frame deletion mutant of a gene encoding a COG4948 protein of Haloferax volcanii was generated. The mutant had a lag phase of 3 days after transition from a richer to a poorer medium, in contrast to the wild-type, and the gene was therefore named “important for transition” (iftA). After inoculation of fresh casamino acids or complex medium with stationary phase wild-type cells, the transcript level of iftA was transiently induced at the onset of growth. In contrast, in minimal (or “poor”) glucose medium, both transcript and protein were present throughout growth, even in late stationary phase. A comparison of the transcriptomes of deletion mutant and wild-type revealed that transcript levels of a very restricted set of genes were differentially regulated, including genes encoding proteins involved in phosphate metabolism, regulators and stress response proteins. Taken together, the results indicate that IftA might have a dual function, i.e., transiently after transition to fresh medium and permanently during growth in glucose medium.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  • Abu-Qarn M, Eichler J (2006) Protein N-glycosylation in Archaea: defining Haloferax volcanii genes involved in S-layer glycoprotein glycosylation. Mol Microbiol 61:511–525

    Article  PubMed  CAS  Google Scholar 

  • Ahmed H, Ettema TJ, Tjaden B, Geerling AC, van der Oost J, Siebers B (2005) The semi-phosphorylative Entner–Doudoroff pathway in hyperthermophilic archaea: a re-evaluation. Biochem J 390:529–540

    Article  PubMed  CAS  Google Scholar 

  • Allers T, Ngo HP, Mevarech M, Lloyd RG (2004) Development of additional selectable markers for the halophilic archaeon Haloferax volcanii based on the leuB and trpA genes. Appl Environ Microbiol 70:943–953

    Article  PubMed  CAS  Google Scholar 

  • Aubert M, Weber E, Gintz B, Chater KF, Decaris B (1997) Inactivation or amplification of the spa2 gene, encoding a potential stationary phase regulator, affects diffferentiation in Streptomyces ambofaciens. Can J Microbiol 43:1118–1125

    Article  CAS  Google Scholar 

  • Babbitt PC et al (1996) The enolase superfamily: a general strategy for enzyme-catalyzed abstraction of the alpha-protons of carboxylic acids. Biochemistry 35:16489–16501

    Article  PubMed  CAS  Google Scholar 

  • Bailey TL, Elkan C (1994) Fitting a mixture model by expectation maximization to discover motifs in biopolymers. Proc Int Conf Intell Syst Mol Biol 2:28–36

    PubMed  CAS  Google Scholar 

  • Baumann A, Lange C, Soppa J (2007) Transcriptome changes and cAMP oscillations in an archaeal cell cycle. BMC Cell Biol 8:21

    Article  PubMed  CAS  Google Scholar 

  • Berstein JA, Kin PH, Cohen SN, Kin-Chao S (2004) Global analysis of Escherichia coli RNA degradosome function using DNA microarrays. Proc Natl Acad Sci USA 101:2758–2763

    Article  CAS  Google Scholar 

  • Bitan-Banin G, Ortenberg R, Mevarech M (2003) Development of a gene knockout system for the halophilic archaeon Haloferax volcanii by use of the pyrE gene. J Bacteriol 185:772–778

    Article  PubMed  CAS  Google Scholar 

  • Brenneis M, Hering O, Lange C, Soppa J (2007) Experimental characterization of cis-acting elements important for translation and transcription in halophilic Archaea. PLoS Genet 3:e229

    Article  PubMed  CAS  Google Scholar 

  • Carpousis AJ (2007) The RNA degradosome of Escherichia coli: an mRNA-degrading machine assembled on RNase E. Annu Rev Microbiol 61:71–87

    Article  PubMed  CAS  Google Scholar 

  • Chomczynski P, Sacchi N (1987) Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem 162:156–159

    Article  PubMed  CAS  Google Scholar 

  • Cline SW, Schalkwyk LC, Doolittle WF (1989) Transformation of the archaebacterium Halobacterium volcanii with genomic DNA. J Bacteriol 171:4987–4991

    PubMed  CAS  Google Scholar 

  • Danner S, Soppa J (1996) Characterization of the distal promoter element of halobacteria in vivo using saturation mutagenesis and selection. Mol Microbiol 19:1265–1276

    Article  PubMed  CAS  Google Scholar 

  • Danson MJ (1988) Archaebacteria: the comparative enzymology of their central metabolic pathways. Adv Microb Physiol 29:165–231

    Article  PubMed  CAS  Google Scholar 

  • Felsenstein J (1996) Inferring phylogenies from protein sequences by parsimony, distance, and likelihood methods. Methods Enzymol 266:418–427

    Article  PubMed  CAS  Google Scholar 

  • Gerlt JA, Babbitt PC, Rayment I (2005) Divergent evolution in the enolase superfamily: the interplay of mechanism and specificity. Arch Biochem Biophys 433:59–70

    Article  PubMed  CAS  Google Scholar 

  • Gregor D, Pfeifer F (2005) In vivo analyses of constitutive and regulated promoters in halophilic archaea. Microbiology 151:25–33

    Article  PubMed  CAS  Google Scholar 

  • Hall TA (1999) BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucl Acids Symp Ser 41:95–98

    CAS  Google Scholar 

  • Herrmann U, Soppa J (2002) Cell cycle-dependent expression of an essential SMC-like protein and dynamic chromosome localization in the archaeon Halobacterium salinarum. Mol Microbiol 46:395–409

    Article  PubMed  CAS  Google Scholar 

  • Huisman GW, Kolter R (1994) Sensing starvation: a homoserine lactone-dependent signaling pathway in Escherichia coli. Science 265:537–539

    Article  PubMed  CAS  Google Scholar 

  • Jolley KA et al (2000) 2-Oxoacid dehydrogenase multienzyme complexes in the halophilic Archaea? Gene sequences and protein structural predictions. Microbiology 146:1061–1069

    PubMed  CAS  Google Scholar 

  • Kapatai G et al (2006) All three chaperonin genes in the archaeon Haloferax volcanii are individually dispensable. Mol Microbiol 61:1583–1597

    Article  PubMed  CAS  Google Scholar 

  • Kim S, Lee SB (2005) Identification and characterization of Sulfolobus solfataricus d-gluconate dehydratase: a key enzyme in the non-phosphorylated Entner–Doudoroff pathway. Biochem J 387:271–280

    Article  PubMed  CAS  Google Scholar 

  • Kirkland PA, Gil MA, Karadzic IM, Maupin-Furlow JA (2007) Genetic and proteomic analyses of a proteasome-activating nucleotidase a mutant of the haloarchaeon Haloferax volcanii. J Bacteriol. doi:10.1128/JB.01196-07)

  • Lamble HJ, Milburn CC, Taylor GL, Hough DW, Danson MJ (2004) Gluconate dehydratase from the promiscuous Entner–Doudoroff pathway in Sulfolobus solfataricus. FEBS Lett 576:133–136

    Article  PubMed  CAS  Google Scholar 

  • Lando JA, Gerlt JA, Kozarich JW, Koo CW, Shah VJ, Kenyon GL, Neidhart DJ, Fujuta S, Petsko GA (1994) The role of lysine 166 in the mechanism of mandelate racemase from Pseudomonas putida: mechanistic and crystallographic evidence for stereospecific alkylation be ®-alpha-phenylglycidate. Biochemistry 33:635–643

    Article  Google Scholar 

  • Lange C et al (2007) Genome-wide analysis of growth phase-dependent translational and transcriptional regulation in halophilic archaea. BMC Genomics 8:415

    Article  PubMed  Google Scholar 

  • Large A et al (2007) Characterization of a tightly controlled promoter of the halophilic archaeon Haloferax volcanii and its use in the analysis of the essential cct1 gene. Mol Microbiol 66:1092–1106

    Google Scholar 

  • Ming YZ, Di X, Gomez-Sanchez EP, Gomez-Sanchez CE (1994) Improved downward capillary transfer for blotting of DNA and RNA. Biotechniques 16:58–59

    PubMed  CAS  Google Scholar 

  • Norais C, Hawkins M, Hartman AL, Eisen JA, Myllykallio H, Allers T (2007) Genetic and physical mapping of DNA replication origins in Haloferax volcanii. PLoS Genet 3:e77

    Article  PubMed  CAS  Google Scholar 

  • Rosenshine I, Zusman T, Werczberger R, Mevarech M (1987) Amplication of specific DNA-sequences correlates with resistance of the archaebacterium halobacterium volcanii to the dihydrofolate reductase inhibitors trimethoprim and methotrexate. Mol Gen Genet 208:518–522

    Article  CAS  Google Scholar 

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

    Google Scholar 

  • Scheuch S, Pfeifer F (2007) GvpD-induced breakdown of the transcriptional activator GvpE of halophilic archaea requires a functional P-loop and an arginine-rich region of GvpD. Microbiology 153:947–958

    Article  PubMed  CAS  Google Scholar 

  • Schneider D, Bruton CJ, Chater KF (1996) Characterization of spaA, a Streptomyces coelicolor gene homologous to a gene involved in sensing starvation in Escherichia coli. Gene 177:243–251

    Article  PubMed  CAS  Google Scholar 

  • Siebers B, Schönheit P (2005) Unusual pathways and enzymes of central carbohydrate metabolism in Archaea. Curr Opin Microbiol 8:695–705

    PubMed  CAS  Google Scholar 

  • Soppa J (2006) From genomes to function: haloarchaea as model organisms. Microbiology 152:585–590

    Article  PubMed  CAS  Google Scholar 

  • Soppa J, Baumann A, Brenneis M, Dambeck M, Hering O, Lange C (2008) Genomics and functional genomics with haloarchaea. Arch Microbiol. doi:10.1007/s00203-008-0376-4

  • Taghbalout A, Rothfield L (2008) RNaseE and RNA helicase B play central roles in the cytoskeletal organization of the RNA degradosome. J Biol Chem (ehead of print). doi:10.1074/jbc.M709118200

  • 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  PubMed  CAS  Google Scholar 

  • van Ooyen J, Soppa J (2007) Three 2-oxoacid dehydrogenase operons in Haloferax volcanii: expression, deletion mutants and evolution. Microbiology 153:3303–3313

    Article  PubMed  CAS  Google Scholar 

  • Wanner C, Soppa J (2002) Functional role for a 2-oxo acid dehydrogenase in the halophilic archaeon Haloferax volcanii. J Bacteriol 184:3114–3121

    Article  PubMed  CAS  Google Scholar 

  • Wessel D, Flugge UI (1984) A method for the quantitative recovery of protein in dilute solution in the presence of detergents and lipids. Anal Biochem 138:141–143

    Article  PubMed  CAS  Google Scholar 

  • Zaigler A, Schuster SC, Soppa J (2003) Construction and usage of a onefold-coverage shotgun DNA microarray to characterize the metabolism of the archaeon Haloferax volcanii. Mol Microbiol 48:1089–1105

    Article  PubMed  CAS  Google Scholar 

  • Zhao A, Gray FC, MacNeill SA (2006) ATP- and NAD+-dependent DNA ligases share an essential function in the halophilic archaeon Haloferax volcanii. Mol Microbiol 59:743–752

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

We are thankful to Thorsten Allers (University of Nottingham, UK) for sharing the H. volcanii strain H26 and the plasmid pTA131. This work was supported by the German Research Foundation (Deutsche Forschungsgemeinschaft) through grant So264/10 in the framework of the Special Research Program SPP1112.

Author information

Authors and Affiliations

  1. Biocentre, Institute for Molecular Biosciences, Goethe-University, Max-von-Laue-Str. 9, 60438, Frankfurt, Germany

    Michael Dambeck & Jörg Soppa

Authors
  1. Michael Dambeck
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jörg Soppa
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Jörg Soppa.

Additional information

Communicated by William Metcalf.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary Table S1 (DOC 43 kb).

Rights and permissions

Reprints and permissions

About this article

Cite this article

Dambeck, M., Soppa, J. Characterization of a Haloferax volcanii member of the enolase superfamily: deletion mutant construction, expression analysis, and transcriptome comparison. Arch Microbiol 190, 341–353 (2008). https://doi.org/10.1007/s00203-008-0379-1

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00203-008-0379-1

Keywords

Search

Navigation