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Highly selective extraction of spiralin from the Spiroplasma citri cell membrane with alkyl-N-sulfobetaines
Abstract:
The extraction of proteins from the membrane of the mollicute (mycoplasma) Spiroplasma citri by sodium N-dodecyl-N,N-dimethyl-3-amino-1-propane sulfonate (SB12) and sodium N-tetradecyl-N,N-dimethyl-3-amino-1-propane sulfonate (SB14) was studied with electrophoretic methods. The membranes were prepared by osmotic lysis of the cells and depleted of the bulk of extrinsic proteins. It was possible to extract up to 35 and 45% of membrane proteins with SB12 and SB14, respectively. Maximal yield was obtained in both cases with detergent concentrations greater than or equal to 5 mumoles/mg of membrane protein. Spiralin, the major protein in the S. citri membrane, was highly selectively solubilized without the loss of antigenicity, with a yield of about 90% with SB12 and close to 100% with SB14, for a detergent concentration greater than or equal to 0.2 M. The degree of selectivity in favour of spiralin was higher with SB12 (purity approximately equal to 70%) than with SB14 (purity approximately equal to 50%). Treatment of the S. citri membrane with high concentrations of SB12 is a simple and fast procedure for partial purification of spiralin. This example shows that, in some cases, it should be possible to modulate the selectivity of the extraction of membrane proteins simply by varying the relative concentration of detergent.
Insights
Sodium N-dodecyl-N,N-dimethyl-3-amino-1-propane sulfonate (SB12) and SB14 effectively extract Spiroplasma citri membrane proteins. SB12 offers a simple method for partial purification of spiralin, the major membrane protein.
Area of Science:
- Membrane protein extraction
- Biochemistry
- Microbiology
Background:
- Spiroplasma citri is a mollicute (mycoplasma) with a distinct membrane protein composition.
- Understanding membrane protein extraction is crucial for studying microbial structures and functions.
Purpose of the Study:
- To investigate the efficacy of two detergents, sodium N-dodecyl-N,N-dimethyl-3-amino-1-propane sulfonate (SB12) and sodium N-tetradecyl-N,N-dimethyl-3-amino-1-propane sulfonate (SB14), for extracting proteins from Spiroplasma citri membranes.
- To evaluate the selectivity of these detergents in solubilizing spiralin, the major membrane protein.
Main Methods:
- Membrane preparation via osmotic lysis of Spiroplasma citri cells.
- Protein extraction using SB12 and SB14 detergents at varying concentrations.
- Electrophoretic analysis to determine protein yield and purity.
Main Results:
- SB12 and SB14 extracted 35% and 45% of membrane proteins, respectively, with maximal yield at detergent concentrations >= 5 µmol/mg.
- Spiralin was selectively solubilized with high yield (90% with SB12, ~100% with SB14) without losing antigenicity at detergent concentrations >= 0.2 M.
- SB12 achieved higher spiralin purity (~70%) compared to SB14 (~50%).
Conclusions:
- SB12 provides a simple, rapid method for partial purification of spiralin from Spiroplasma citri membranes.
- Detergent concentration can be modulated to control the selectivity of membrane protein extraction.