ORNITHINE DECARBOXYLASE MODIFICATION AND INACTIVATION

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The polyamines are essential components of the eucaryotic cell. Spermidine and spermine levels frequently correlate with growth rate and are elevated in rapidly growing embryonic and neoplastic tissues. The activity of the rate-limiting enzymes in the polyamine biosynthetic pathway, ornithine decarboxylase (ODC), changes quickly in response to many different variations in growth conditions. Athough some control of the activity of this enzyme is exerted at the level of synthesis, this enzyme is most unusual in that it has a turnover rate far greater than any other eucaryotic enzyme, allowing extremely rapid modifications in this enzyme activity. Very little is known, however, about this very efficient, specific enzyme inactivation mechanism or its control. Our recent studies suggest that this rapid enzyme turnover can be separated into at least two phases. First there is a polyamine-dependent step in which the charge characteristics of the enzyme are modified slightly, creating isolatable alternate forms of the enzyme monomer. This is followed by an energy-dependent inactivation of the modified enzyme leading to eventual degradation. We intend to investigate ornithine decarboxylase turnover and its control in vivo, the polyamine-dependent modification of this enzyme and the mechanism of its subsequent energy-dependent inactivation. Our goal is to reveal the mechanisms and control of this specific, and tremendously effective, enzyme deactivation. We will use a rat hepatoma cell line (HTC) and its subline (HMOA) which is deficient at some point in this ODC turnover. In vivo studies will involve controlled enzyme stabilization by use of inhibitors of spermidine synthesis such as methylglyoxal bis (guanylhydrazone) and dicyclohexylamine, as well as metabolic energy inhibitors. Proposed in vitro studies include isolation and complete characterization of the energy-dependent ODC inactivation. In addition to elucidating this facet of the control of a most enigmatic enzyme, these data should specifically help us understand the increase in ODC activity seen in response to carcinogens and viral transformation of normal cells that appear to be related to abnormalities in this enzyme turnover.