GLYCINE PROTECTS AGAINST STRONG PROTEIN-DENATURING ACTIVITY OF OLEUROPEIN - A PHENOLIC COMPOUND IN PRIVET LEAVES

In previous studies, we reported the existence of a high concentration of free glycine in the digestive juice of several Lepidoptera larvae, particularly in the digestive juice of species that feed on the prive t tree, Ligustrum obtusifolium. The water extract of privet leaves sho wed very strong protein-denaturing activity and lysine-decreasing acti vity, which closely resembled activity of oxidized polyphenolics. Addi tion of 1% glycine to the extract could completely inhibit these activ ities. Free glycine may be secreted into the digestive juice by larvae as an adaptive mechanism for chemical defense against its host plants . The protein-denaturing compound in priver leaves is present in the c ytosol or in the vacuoles of the leaf cells. The compound does not sho w protein-denaturing activity without oxidation, but when mixed with i ntact organelles under low osmotic conditions to give an osmotic shock , a very high protein-denaturing activity is produced. Our results sug gest that the privet tree is endowed with a defense mechanism in which a stable compound in the cytosol or in the vacuoles is activated into a chemically active denaturant by an enzyme present in the organelles (including chloroplasts) after the leaves are eaten by insects and th e organelles are broken by osmotic shock or by digestive mechanisms. B ased upon HPLC and NMR data, we conclude that the denaturing compound is cleuropein, an o-dihydroxyphenolic compound. This compound makes up 3% of the wet weight of privet leaves. The protein-denaturing activit y of purified oleuropein activated by the leaf enzyme is high enough t o account for all the denaturing activity in a water extract of privet leaves. The denaturing reaction is completely inhibited by free glyci ne. Our results suggest that the protein-denaturing activity and lysin e-decreasing activity of privet leaves are caused by oxidized polyphen olics, and that some insects secrete free glycine to counter the denat uring activity of oxidized phenolics. The chemical mechanism of counte raction by glycine is also considered. Free glycine in the midgut of i nsects probably protects proteins from denaturation by competing with proteins for oxidized phenolics.