Over-expression, purification and phosphorylation of the Bacillus subtilis CheA protein.
The CheA (Bacillus subtilis) protein was over-expressed in a CheA minus E. Coli strain. Cells induced for the over-expression of CheA were lysed by passage through a French press device. The French press lysate was partially purified by ammonium sulfate precipitation and the resulting lysate was loaded onto a cibacron blue column and eluted with a NaCl gradient. CheA was finally purified by separation with a G-150 column. The purity and identity of CheA was verified by SDS-PAGE, CnBr digestion and amino acid analysis. Purified material autophosphorylated at the expected molecular weight of CheA.
Fuhrer
D K
DK
Department of Medicine (Hematology/Oncology), Indiana University School of Medicine, Indianapolis 46202, USA.
D000596:Amino Acids / Q000032:analysis; D001412:Bacillus subtilis / Q000737:chemistry*; D001426:Bacterial Proteins; D011232:Chemical Precipitation; D002845:Chromatography / Q000379:methods; D003488:Cyanogen Bromide / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D008565:Membrane Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D000072236:Methyl-Accepting Chemotaxis Proteins; D008970:Molecular Weight; D010766:Phosphorylation
D000596:Amino Acids; D001426:Bacterial Proteins; D008565:Membrane Proteins; D000072236:Methyl-Accepting Chemotaxis Proteins; D003488:Cyanogen Bromide