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Mycolylarabinogalactan Peptidoglycan
Reagent:
Mycolylarabinogalactan Peptidoglycan Complex, mAGP, Cell Wall Core
Default Quantity:
5 mg
Production system:
Each strain is grown to late-log phase (day 14) in glycerol-alanine-salts (GAS) medium, washed with PBS pH 7.4 and inactivated by gamma-irradiation. The bacilli are suspended (2 g/ml) in PBS containing DNase, and RNase, and broken in a French Press pressure cell at 4°C. Unbroken cells are removed by low speed (3,000 xg) centrifugation. The cell wall is isolated by centrifugation at 27,000 xg for one hour , the pellet suspended in PBS with 2% SDS and the cell wall associated proteins and lipoglycans are extracted at room temperature. Residual contaminating proteins are removed from the cell wall by further SDS extraction and proteinase-K treatment at 55°C, followed by several extractions with boiling SDS. The resulting mAGP is washed several times with water, and residual SDS is removed by extraction with large volumes of acetone.
Notes:
The individual products that constitute the cell wall core (arabinogalatan, mycolic acid and peptidoglycan) are also available for specific research needs. QC of the mAGP includes quantification of protein and SDS contamination.
References:
Daffe M., et al. J. Biol. Chem. 265:6734, 1990.
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