Abstract
Abstract Cyclic nucleotide phosphodiesterase has been prepared from the liver, skeletal muscle, heart muscle, kidney, epididymal fat pad, and brain cortex of the rat. Kinetic analysis indicated that all of these tissues, with the exception of liver exhibit complex behavior indicative of multiple enzymatic activities. The phosphodiesterase activities of brain cortex, kidney, and adipose tissue have been resolved on Bio-Gel A-1.5m gel filtration columns into three fractions. These are an excluded fraction, a high molecular weight fraction in which the enzymatic affinity for cyclic guanosine 3',5'-monophosphate is greater than that for cyclic adenosine 3',5'-monophosphate, and a lower molecular weight fraction with a high affinity for cyclic adenosine 3',5'-monophosphate. The lower molecular weight enzyme appears to be negatively cooperative. The muscle preparations have little, if any, of the excluded fraction which in kidney, adipose tissue, and brain has kinetic behavior similar to that of the lower molecular weight enzyme. Liver has no detectable phosphodiesterase activity in any except the intermediate fraction which has a molecular weight of approximately 400,000. This enzymatic activity has been purified 88-fold by centrifugation, ammonium sulfate fractionation, and gel filtration. The potential for metabolic control through regulation of cyclic nucleotide phosphodiesterase is discussed in view of the existence of at least two distinct enzymatic activities in all of tissues studied except the liver.
Highlights
A detailed discussion of the interpretation of this kinetic behavior has been previously presented in relation to the brain cortex enzymes
Kidney-Agarose gel filtration demonstrates the existence of Fraction I, II, and III enzymes in kidney (Fig. 1) analogous to the profiles obtained for brain preparations
Adipose Tissue-Agarose A-1.5m gel fractionation profiles of the sonically disrupted infranatant from epididymal fat pad and for the enzyme prepared from isolated fat cells are similar
Summary
Cyclic nucleotide phosphodiesterase has been prepared from the liver, skeletal muscle, heart muscle, kidney, epididymal fat pad, and brain cortex of the rat. The phosphodiesterase activities of brain cortex, kidney, and adipose tissue have been resolved on Bio-Gel A-1.5m gel filtration columns into three fractions These are an excluded fraction, a high molecular weight fraction in which the enzymatic affinity for cyclic guanosine 3’,5’-monophosphate is greater than that for cyclic adenosine 3’,5’monophosphate, and a lower molecular weight fraction with a high affinity for cyclic adenosine 3’,5’-monophosphate. We have reported the existence of multiple forms of cyclic nucleotide phosphodiesterases in rat brain cortex [1] These enzymes can be differentiated by separation based on molecular size and by their distinct affinities for cyclic AMP’ and cyclic GMP. This report presents a physical separation and kinetic analysis of this enzyme system for some other tissues of the rat including heart muscle, skeletal muscle, kidney, adipose tissue, and liver This information leads to some interesting possibilities in the regulation of cyclic nucleotide levels. Fraction I will denote the membrane bound enzyme with a high affinity for cyclic AMP, Fraction II will denote the separated higher molecular weight enzyme with an affinity for cyclic GMP greater than its affinity for cyclic AMP, and Fraction III will denote the separated lower molecular weight enzyme which is kinetically similar to Fraction I
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