PROPERTIES OF CLOCK-CONTROLLED AND CONSTITUTIVE N-ACETYLTRANSFERASES FROM CHICK PINEAL CELLS

The pineal gland synthesizes its hormone melatonin (O-methyl-N-acetyls erotonin) from serotonin. Acetyl-CoA: serotonin N-acetyltransferase (S NAT), the enzyme that catalyzes the committed step in this biosynthesi s, is largely restricted to the pineal gland and is regulated by adren ergic and circadian mechanisms. Another enzyme, acetyl-CoA: arylamine N-acetyltransferase (ANAT), having an apparently similar activity, is also present in the pineal. This enzyme, however, is not rhythmically regulated. SNAT activity of cultured chick pineal cells was obtained w ithout ANAT after ammonium sulfate precipitation. ANAT activity was re tained without SNAT activity after pre-incubation at 37 degrees C. Thu s, each enzyme could be examined independently. Overlap in substrate s pecificity between the two enzymes was minimal. Kinetic analysis of th e separated enzyme activities revealed that while SNAT operates via a random or ordered bi bi mechanism, ANAT catalysis occurs through a pin g pong bi bi mechanism with substrate inhibition by acetyl-CoA. By siz e-exclusion chromatography, ANAT was confirmed to be 30-35 kDa, and SN AT was estimated at 15-20 kDa. Taken together, these results indicate that the two enzymes differ in their structure, reactivity, stability, and mechanism of catalysis.