The stereochemical course of phospho group transfer catalyzed by rat liver 6-phosphofructo-2-kinase.

Adenosine [gamma-(S)-16O,17O,18O]triphosphate was used as substrate in the phosphokinase reaction catalyzed by 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase. The product D-fructose 2,6-[2-16O,17O,18O]bisphosphate was then used as substrate in the alkaline phosphatase-mediated transfer of the phospho groups to (S)-butane-1,3-diol, where the configuration at phosphorus has been determined. Although there was approximately equal transfer by alkaline phosphatase of the labeled 2- and the unlabeled 6-phospho groups, the subsequent assignment of configuration of the chiral phospho group from the 2-position was unambiguous.

Hepatic 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase: phosphate dependence and effects of other oxyanions.

The effects of various oxyanions on the activities of rat liver 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (EC 2.7.1.

The effect of arabinose 1,5-bisphosphate on rat hepatic 6-phosphofructo-1-kinase and fructose-1,6-bisphosphatase.

The alpha- and beta-anomers of arabinose 1,5-bisphosphate and ribose 1,5-bisphosphate were tested as effectors of rat liver 6-phosphofructo-1-kinase and fructose-1,6-bisphosphatase. Both anomers of arabinose 1,5-bisphosphate activated the kinase and inhibited the bisphosphatase. The alpha-anomer was the more effective kinase activator while the beta-anomer was the more potent inhibitor of the bisphosphatase.

Isolation and characterization of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase from bovine liver.

6-Phosphofructo-2-kinase and fructose-2,6-bisphosphatase activities were copurified to homogeneity from bovine liver. The purification scheme consisted of polyethylene glycol precipitation, anion-exchange and Blue-Sepharose chromatography, substrate elution from phosphocellulose, and gel filtration. The bifunctional enzyme had an apparent molecular weight of 102,000 and consisted of two subunits (Mr 49,000).

Amino acid sequence of the phosphorylation site of rat liver 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase.

6-Phosphofructo-2-kinase/fructose-2,6-bisphosphatase from rat liver was phosphorylated by cyclic AMP-dependent protein kinase and [gamma-32P]ATP. Treatment of the 32P-labeled enzyme with thermolysin removed all of the radioactivity from the enzyme core and produced a single labeled peptide. The phosphopeptide was purified by ion exchange chromatography, gel filtration, and reverse phase high pressure liquid chromatography.