Omega-aminoalkyl agaroses in the resolution of enzymes involved in regulation of glutamine metabolism.
Systematic examination of a homologous series of omega-aminoalkyl agaroses showed that the pentyl derivative (Seph-C5-NH2) was best suited for the retention and subsequent separation of several proteins involved in the regulation of glutamine metabolism in Escherichia coli, including: glutamine synthetase [EC 6.3.1.2; L-glutamate:ammonia ligase (ADP-forming)], ATP:glutamine synthetase adenylyltransferase (EC 2.7.7.42), the PII regulatory protein which regulates the adenylylation and deadenylylation activities of the adenylyltransferase, the UTP:PII protein uridylyltransferase, and the uridylyl removing enzyme which catalyzes the removal of uridylyl groups from uridylylated PII protein. Resolution of these proteins was achieved by gradually increasing the concentration of KCl in the eluant, which resulted in consecutive detachment of the proteins from the column. Proteins that co-elute from a DEAE-cellulose column can be resolved and further purified on epsilon-aminopentyl agarose, probably due to the fact that with the homologous series it is possible to adjust the contribution of hydrophobic interactions for optimal resolution.
D001276:ATP Phosphoribosyltransferase / Q000302:isolation & purification; D001426:Bacterial Proteins / Q000302:isolation & purification; D002845:Chromatography / Q000379:methods*; D004926:Escherichia coli / Q000201:enzymology*; D005974:Glutamate-Ammonia Ligase / Q000302:isolation & purification*; D011134:Polysaccharides / Q000031:analogs & derivatives*; D012685:Sepharose / Q000031:analogs & derivatives*
D001426:Bacterial Proteins; D011134:Polysaccharides; D012685:Sepharose; D001276:ATP Phosphoribosyltransferase; D005974:Glutamate-Ammonia Ligase