This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Wilson, H. L.
Right arrow Articles by Maupin-Furlow, J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Wilson, H. L.
Right arrow Articles by Maupin-Furlow, J.

 Previous Article  |  Next Article 

Journal of Bacteriology, March 2000, p. 1680-1692, Vol. 182, No. 6
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Biochemical and Physical Properties of the Methanococcus jannaschii 20S Proteasome and PAN, a Homolog of the ATPase (Rpt) Subunits of the Eucaryal 26S Proteasomedagger

Heather L. Wilson, Mark S. Ou, Henry C. Aldrich, and Julie Maupin-Furlow*

Department of Microbiology and Cell Science, University of Florida, Gainesville, Florida 32611-0700

Received 17 September 1999/Accepted 3 January 2000

The 20S proteasome is a self-compartmentalized protease which degrades unfolded polypeptides and has been purified from eucaryotes, gram-positive actinomycetes, and archaea. Energy-dependent complexes, such as the 19S cap of the eucaryal 26S proteasome, are assumed to be responsible for the recognition and/or unfolding of substrate proteins which are then translocated into the central chamber of the 20S proteasome and hydrolyzed to polypeptide products of 3 to 30 residues. All archaeal genomes which have been sequenced are predicted to encode proteins with up to ~50% identity to the six ATPase subunits of the 19S cap. In this study, one of these archaeal homologs which has been named PAN for proteasome-activating nucleotidase was characterized from the hyperthermophile Methanococcus jannaschii. In addition, the M. jannaschii 20S proteasome was purified as a 700-kDa complex by in vitro assembly of the alpha  and beta  subunits and has an unusually high rate of peptide and unfolded-polypeptide hydrolysis at 100°C. The 550-kDa PAN complex was required for CTP- or ATP-dependent degradation of beta -casein by archaeal 20S proteasomes. A 500-kDa complex of PAN(Delta 1-73), which has a deletion of residues 1 to 73 of the deduced protein and disrupts the predicted N-terminal coiled-coil, also facilitated this energy-dependent proteolysis. However, this deletion increased the types of nucleotides hydrolyzed to include not only ATP and CTP but also ITP, GTP, TTP, and UTP. The temperature optimum for nucleotide (ATP) hydrolysis was reduced from 80°C for the full-length protein to 65°C for PAN(Delta 1-73). Both PAN protein complexes were stable in the absence of ATP and were inhibited by N-ethylmaleimide and p-chloromercuriphenyl-sulfonic acid. Kinetic analysis reveals that the PAN protein has a relatively high Vmax for ATP and CTP hydrolysis of 3.5 and 5.8 µmol of Pi per min per mg of protein as well as a relatively low affinity for CTP and ATP with Km values of 307 and 497 µM compared to other proteins of the AAA family. Based on electron micrographs, PAN and PAN(Delta 1-73) apparently associate with the ends of the 20S proteasome cylinder. These results suggest that the M. jannaschii as well as related archaeal 20S proteasomes require a nucleotidase complex such as PAN to mediate the energy-dependent hydrolysis of folded-substrate proteins and that the N-terminal 73 amino acid residues of PAN are not absolutely required for this reaction.

* Corresponding author. Mailing address: Department of Microbiology and Cell Science, University of Florida, Gainesville, FL 32611-0700. Phone: (352) 392-4095. Fax: (352) 392-5922. E-mail: jmaupin{at}ufl.edu.

dagger Journal series R07342 of the Florida Agricultural Experiment Station.

Journal of Bacteriology, March 2000, p. 1680-1692, Vol. 182, No. 6
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


This article has been cited by other articles:

  • Horwitz, A. A., Navon, A., Groll, M., Smith, D. M., Reis, C., Goldberg, A. L. (2007). ATP-induced Structural Transitions in PAN, the Proteasome-regulatory ATPase Complex in Archaea. J. Biol. Chem. 282: 22921-22929 [Abstract] [Full Text]  
  • Sharon, M., Witt, S., Glasmacher, E., Baumeister, W., Robinson, C. V. (2007). Mass Spectrometry Reveals the Missing Links in the Assembly Pathway of the Bacterial 20 S Proteasome. J. Biol. Chem. 282: 18448-18457 [Abstract] [Full Text]  
  • Madding, L. S., Michel, J. K., Shockley, K. R., Conners, S. B., Epting, K. L., Johnson, M. R., Kelly, R. M. (2007). Role of the {beta}1 Subunit in the Function and Stability of the 20S Proteasome in the Hyperthermophilic Archaeon Pyrococcus furiosus. J. Bacteriol. 189: 583-590 [Abstract] [Full Text]  
  • Farhoud, M. H., Wessels, H. J. C. T., Steenbakkers, P. J. M., Mattijssen, S., Wevers, R. A., van Engelen, B. G., Jetten, M. S. M., Smeitink, J. A., van den Heuvel, L. P., Keltjens, J. T. (2005). Protein Complexes in the Archaeon Methanothermobacter thermautotrophicus Analyzed by Blue Native/SDS-PAGE and Mass Spectrometry. Mol. Cell. Proteomics 4: 1653-1663 [Abstract] [Full Text]  
  • Reuter, C. J., Kaczowka, S. J., Maupin-Furlow, J. A. (2004). Differential Regulation of the PanA and PanB Proteasome-Activating Nucleotidase and 20S Proteasomal Proteins of the Haloarchaeon Haloferax volcanii. J. Bacteriol. 186: 7763-7772 [Abstract] [Full Text]  
  • Kaczowka, S. J., Maupin-Furlow, J. A. (2003). Subunit Topology of Two 20S Proteasomes from Haloferax volcanii. J. Bacteriol. 185: 165-174 [Abstract] [Full Text]  
  • Koonin, E. V., Wolf, Y. I., Aravind, L. (2001). Prediction of the Archaeal Exosome and Its Connections with the Proteasome and the Translation and Transcription Machineries by a Comparative-Genomic Approach. Genome Res 11: 240-252 [Abstract] [Full Text]  
  • Franzetti, B., Schoehn, G., Ebel, C., Gagnon, J., Ruigrok, R. W. H., Zaccai, G. (2001). Characterization of a Novel Complex from Halophilic Archaebacteria, Which Displays Chaperone-like Activities in Vitro. J. Biol. Chem. 276: 29906-29914 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»