Human prolyl hydroxylase. Purification, radioimmunoassay and clinical studies in liver diseases

Abstract

Prolyl hydroxylase was purified from human placentae, specific antiserum against it was prepared, and a new radioimmunoassay system employing 125I-labelled enzyme preparation was established. The molecular weight of the placental enzyme was shown to be 320,000 by gel filtration. SDS-polyacrylamide gel electrophoresis showed two bands of unequal intensity having molecular weights of 60,000 and 130,000. Their amino acid compositions were identical to each other, suggesting the polypeptide with a molecular weight of 130,000 might be a dimer of the polypeptide with a molecular weight of 60,000. The new radioimmunoassay established had a sensitivity of the order of 10 ng/ml, indicating it was more sensitive than previous radioimmunoassay employing 3H-labelling method. Clinical studies on patients with liver diseases disclosed that the concentrations of serum immunoreactive prolyl hydroxylase were elevated both in cases of hepatocellular damage and in cases of cholestasis. In cases of hepatocellular damage the enzyme behaved like cytoplasmic enzymes such as glutamic oxaloacetic transaminase, glutamic pyruvic transaminase and lactic dehydrogenase, but in cases of cholestasis it resembled biliary enzymes such as alkaline phosphatase and gamma-glutamyl transpeptidase. This result might be associated with the peculiar location of the enzyme within the cell, in the membrane of rough endoplasmic reticulum.

Are serum prolyl 4-hydroxylase useful as noninvasive markers of liver fibrosis in patients with biliary atresia?

“…Its concentration in serum is considered to selectively reflect hepatic necrosis because it is only released into the circulation when a cell is destroyed [3]. Thus, when there is hepatocellular damage there is concurrent elevation of serum PH and cytoplasmic enzymes, and when there is cholestasis there is concurrent elevation of serum PH with biliary enzymes, suggesting that a quantitative assay for PH can be used to determine the amount of enzyme released from the liver into the bloodstream [4]. In other words, serum PH is a reliable noninvasive marker for hepatic fibrogenesis and fibrosis.…”

“…Several papers have reported that the release of PH into serum in response to hepatocellular damage and cholestatic diseases is related to the fact that it is localized in RER membranes-that is, when there is hepatocellular damage, PH may detach from RER membranes and be released into the blood with other cytoplasmic enzymes, and in cholestasis, PH could increase in the blood by a mechanism similar to that which controls the release of biliary enzymes [4].…”