[Isolation and properties of glycogen phosphorylase b from rabbit liver].
Purification of dephosphorylated form of rabbit liver glycogen phosphorylase (phosphorylase b) has been carried out. The purification procedure included 3 steps:sedimentation of protein-glycogen pellet by centrifugation at 62 000 x g during 2 hours, chromathography on omega-amino-hexyl-Sepharose column and isoelectric focusing. The yield of the enzyme is about 60%, the specific activity at glucose-1-phosphate concentration 100mM is 35 mkmol of Pi/mg protein-min. The final preparation revealed apparent homogeneity during ultracentrifugation and polyacrylamide gel electrophoresis. The subunit molecular weight of 100 000 was determined by disc-electrophoresis in the presence of sodium dodecyl sulfate. The absorption spectrum of liver phosphorylase b shows a protein maximum at 280 nm and a second peak at 333 nm produced by bound PLP. A kinetic analysis of initial rates with variable concentrations of the substrate revealed the homotropic cooperativity of glucose-1-phosphate binding sites. This cooperative effect is more pronounced in the presence of inhibitors, glucose and ATP and less pronounced in the presence of AMP.
D000249:Adenosine Monophosphate / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D001665:Binding Sites; D005947:Glucose / Q000494:pharmacology; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008970:Molecular Weight; D006005:Phosphorylases / Q000302:isolation & purification* / Q000378:metabolism; D011817:Rabbits; D013329:Structure-Activity Relationship
D000249:Adenosine Monophosphate; D000255:Adenosine Triphosphate; D006005:Phosphorylases; D005947:Glucose
Vydelenie glikogenfosforilazy b iz pecheni krolika i issledovanie nekotorykh svoistv fermenta