Rapid purification of mitochondrial hexokinase from rat brain by a single affinity chromatography step on Affi-Gel blue.
The mitochondrial hexokinase from rat brain, selectively released from mitochondria by the action of glucose 6-phosphate, can be purified to greater than 90% homogeneity by a single affinity chromatography step on Affi-Gel Blue; the Cibacron Blue F3GA ligand bound to this matrix serves as an analog of ATP, the normal substrate for the enzyme, and selective elution is accomplished using glucose 6-phosphate which is a competitive ligand vs. ATP. With this and other modifications to the previously described procedure highly purified enzyme is readily obtained in good yield and with retention of the ability to rebind to mitochondria.
Wilson
J E
JE
Biochemistry Dept., Michigan State University, East Lansing 48823.
D000818:Animals; D001923:Brain Chemistry; D002846:Chromatography, Affinity; D003720:Densitometry; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D005958:Glucosephosphates / Q000378:metabolism; D006593:Hexokinase / Q000302:isolation & purification*; D008297:Male; D008928:Mitochondria / Q000201:enzymology*; D051381:Rats; D011919:Rats, Inbred Strains; D013379:Substrate Specificity
D005958:Glucosephosphates; D006593:Hexokinase