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Expression and Purification of Mitochondrial RNA Polymerase and Transcription Factor A from Drosophila melanogaster

  • John P. Gajewski
  • Jamie J. Arnold
  • Tiina S. Salminen
  • Laurie S. Kaguni
  • Craig E. CameronEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1351)

Abstract

Mitochondrial gene expression is essential in all organisms. Our understanding of mitochondrial transcription on a biochemical level has been limited by the inability to purify the individual protein components involved in mitochondrial gene expression. Recently, new systems have been identified that permit purification of these proteins from bacteria. However, the generalizability of these systems is not clear. Here, we have applied the technology from the Cameron lab to express and purify mitochondrial RNA polymerase and transcription factor A from Drosophila melanogaster. We show that the use of SUMO system to produce SUMO fusion proteins in bacteria is effective not only for the human and mouse proteins, but also for the fly proteins. The application of this system to produce the mitochondrial proteins from other organisms should permit detailed understanding of mitochondrial transcription from any organism.

Key words

Mitochondria Mitochondrial transcription Transcription factor A Mitochondrial RNApolymerase Protein expression Protein purification 

Notes

Acknowledgments

L.S.K. was supported by the NIH grant 45295 and a Finland Distinguished Professor Programme grant from the Academy of Finland. T.S.S. was supported by the Academy of Finland.

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Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • John P. Gajewski
    • 1
  • Jamie J. Arnold
    • 1
  • Tiina S. Salminen
    • 2
  • Laurie S. Kaguni
    • 2
    • 3
  • Craig E. Cameron
    • 1
    Email author
  1. 1.Department of Biochemistry and Molecular BiologyThe Pennsylvania State UniversityUniversity ParkUSA
  2. 2.Institute of Biosciences and Medical TechnologyUniversity of TampereTampereFinland
  3. 3.Department of Biochemistry and Molecular Biology, Center for Mitochondrial Science and MedicineMichigan State UniversityEast LansingUSA

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