Abstract

The effect of several biogenic amines that are substrates for adrenal phenylethanolamine N-methyltransferase (PNMT) on thermal and tryptic stability of the enzyme is described. While all substrates tested were somewhat effective in stabilizing the enzyme against tryptic degradation, only phenylethanolamine, phenylethylamine and octopamine were effective in stabilizing PNMT against thermal denaturation. Stabilization of PNMT was maximal when the m and p positions of the phenylethylamine nucleus were unsubstituted. Thus phenylethylamine was the most effective stabilizing compound studied, while norepinephrine was least effective. Epinephrine was completely ineffective in stabilizing the enzyme against proteolytic or thermal breakdown. The interaction of S-adenosylmethionine and substrate in protecting PNMT against thermal and tryptic degradation was also tested. While both substrate and S-adenosylmethionine protect the enzyme against tryptic proteolysis when present singly, the combination of the two compounds affords a degree of stabilization that is substantially greater than that seen by either compound alone. Moreover, the presence of substrate increases the affinity of the enzyme for S-adenosylmethionine, thus reducing the concentration of S-adenosylmethionine required for maximal stabilization of the enzyme. S-adenosylmethionine does not increase the affinity of PNMT for norepinephrine, nor does it augment the stabilization afforded by substrate. These results suggest that in vivo PNMT proteolysis is controlled by the constituents of the PNMT reaction, particularly S-adenosylmethionine. We postulate that interaction of the transferase with its substrate and with S-adenosylmethionine alters the spatial conformation of the protein in such a way that proteolytically vulnerable sites on the molecule are protected.