LIVER PHOSPHORYLASE-B KINASE - CYCLIC-AMP-MEDIATED ACTIVATION AND PROPERTIES OF THE PARTIALLY PURIFIED RAT-LIVER ENZYME

Phosphorylase b kinase was extensively purified from rat liver. It was isolated in a form which could be activated 20–30‐fold by a preincubation with adenosine 3′ : 5′‐monophosphate (cyclic AMP) and ATP‐Mg. This activation was time‐dependent, and was paralleled by a simultaneous incorporation of 32P from [γ‐32P]ATP into two polypeptides which comigrated in sodium dodecyl sulfate gel electrophoresis with the α and β subunits of rabbit skeletal muscle phosphorylase b kinase. The liver enzyme was eluted from Sepharose 4B and Bio‐Gel A‐50m columns at the same place as muscle phosphorylase b kinase, which is indicative of a molecular weight of 1.3 × 106. After activation, the most purified liver preparation had a specific activity about 10‐fold less than the homogeneous muscle enzyme at pH 8.2. The inactive enzyme form had a pronounced pH optimum around pH 6.0, whereas the activated form was mostly activeabove neutral pH. The activation of the enzyme reduced the Km for its substrate phosphorylase b severalfold. Liver phosphorylase b kinase was shown to be partially dependent on Ca2+ ions for its activity: addition of 0.5 mM [ethylenebis‐(oxoethylenenitrilo)]tetraacetic acid (EGTA) to the phosphorylase b kinase assay increased the Km for phosphorylase b about twofold for both the inactive and the activated form of liver phosphorylase b kinase, but affected the V of the inactive species only. Copyright © 1979, Wiley Blackwell. All rights reserved