FIG. 1.

Irreversible inactivation of ODC by cell extracts. (A) ODC metabolically labeled with [³⁵S]methionine in HTC cells was incubated with or without cell extracts that had been supplemented with AZ and ATP. Irreversible inactivation and degradation of ODC were determined as described in Materials and Methods. (B) ODC (a mixture of in vitro-translated labeled rat ODC and cold ODC purified from rat liver) was incubated with cell extracts in an inactivation/degradation reaction mixture. The assay was the same as for Fig. 1A, except that AZ was withdrawn or Z-LLL-CHO (100 μM) was added. When the effect of clasto-lactacystin β-lactone was examined, cell extracts were preincubated with the inhibitor (200 μM) at 37°C for 15 min in an inactivation/degradation reaction mixture without ODC, and then a one-sixth volume of ODC was added. The inactivation and degradation of ODC are represented by filled and open columns, respectively. Results are shown as percentages of the values obtained in the complete reaction mixture without inhibitors.