The assay of Met-enkephalin aminopeptidase with [125I]Met-enkephalin

Presented here are procedural modifications which permit the utilization of 125I-labeled Met-enkephalin as substrate in the assay of rat brain enkephalin aminopeptidase. Th hydrolysis of enkephalin is monitored by the release of [125I]tyrosine separated on Porapak Q. The release of tyrosine is proportionate with both increasing time and tissue concentration. The estimated Km is near 10(-4) M and the enzyme activity can be inhibited more than 95% with puromycin. The majority of the enzyme activity remains in the 100,000 x g supernatant following differential centrifugation.