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Separation of ribosomal protein-protein crosslinks fromBacillus stearothermophilus and ribosomal proteins from archaebacteria by high performance liquid chromatography

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Summary

High performance liquid chromatography was used to separate proteins derived from ribosomes of the archaebacteriumMethanococcus vannielii. Several methods of separation were tested: size exclusion chromatography, reversed phase and ion-exchange chromatography. Best results were obtained using reversed phase columns and volatile buffers that allow direct sequence analysis of the proteins.

In addition, HPLC was used to separate crosslinked protein-protein pairs fromBacillus stearothermophilus ribosomes on a preparative scale. Identification of crosslinked amino acids is a prerequisite for more detailed topographical investigations of this cell organelle. The purification of the 50S crosslink L23–L29 was achieved by a combination of two different reversed phase columns, and the 30S crosslink S13–S19 was purified after salt extraction by size exclusion chromatography.

The methods described here allow a rapid preparation of ribosomal proteins for structural and functional investigations.

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Abbreviations

HPLC:

High Performance Liquid Chromatography

M. vannielii :

Methanococcus vannielii

B. stearothermophilus :

Bacillus stearothermophilus

2D:

two-dimensional

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Authors and Affiliations

  1. Abteilung Wittmann, Max-Planck-Institut für Molekulare Genetik, Ihnestr. 63-73, D-1000, Berlin 33, FRG

    Roza Maria Kamp, J. Brockmöller & Brigitte Wittmann-Liebold

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  1. Roza Maria Kamp
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  2. J. Brockmöller
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  3. Brigitte Wittmann-Liebold
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Kamp, R.M., Brockmöller, J. & Wittmann-Liebold, B. Separation of ribosomal protein-protein crosslinks fromBacillus stearothermophilus and ribosomal proteins from archaebacteria by high performance liquid chromatography. Chromatographia 22, 249–254 (1986). https://doi.org/10.1007/BF02268768

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