Summary
High performance liquid chromatography was used for the purification of ribosomal proteins derived from the halophilic archaebacteriumHalobacterium marismortui. Separation was performed using size exclusion chromatography, reversed phase and ion-exchange chromatography. For reversed phase separation several solvent systems were te sted including isopropanol and acetonitrile. Tris/citrate buffer containing 30% N,N-dimethylformanide was employed in combination with a DEAE-anoin-exchanger.
The influence of the protein extraction method, e.g. lithium chloride versus acetic acid extraction, on the resolution of this highly acidic proteion mixture was investigated and the recovery of single proteins estimaetd. Best results were obtained using the acetic acid extract for reversed phase chromatography. This method lead to the purification of 40% of the proteins from the 50S subunit in one HPLC run. Some of the purified proteins whose sequence was not yet known were subjected to N-termnal and further sequence analysis in order to complete our data of ribsomal proteins from this archaebacterium. The knowledge of the halobacterial ribosomal protein sequences will facilitate further structural and evolutionary studies.
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Abbreviations
- TP30:
-
total ribsomal proteins from 30S subunits
- TP50:
-
total ribosomal proteins from 50S subunits
- TP50LiCl:
-
total ribsomal proteins from 50S subunits extracted with lithium chloride
- TP50AcOH:
-
total ribosmal proteins from 50S subunits extracted with acetic acid
- H. marismortui :
-
Halobacterium marismortui
- HPLC:
-
High-Performance Liquid Chromatography
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Bergmann, U., Wittmann-Liebold, B. Purification of ribosomal proteins from the extreme halophilic archaebacteriumHalobacterium marismortui by high-performance liquid chromatography. Chromatographia 30, 707–712 (1990). https://doi.org/10.1007/BF02269748
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DOI: https://doi.org/10.1007/BF02269748