Role of the nonheme Fe(II) center in the biosynthesis of the plant hormoneethylene

The final step of ethylene biosynthesis in plants is catalyzed by the enzym e 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase (ACCO). In addition t o ACC, Fe(II), O-2, CO2, and ascorbate are required for in vitro enzyme act ivity. Direct evidence for the role of the Fe(II) center in the recombinant avocado ACCO has now been obtained through formation of enzyme (substrate or cofactor) NO complexes. These NO adducts convert the normally EPR-silent ACCO complexes into EPR-active species with structural properties similar to those of the corresponding O-2 complexes. It is shown here that the tern ary Fe(II)ACCO . ACC . NO complex is readily formed, but no Fe(II)ACCO . as corbate . NO complex could be observed, suggesting that ascorbate and NO ar e mutually exclusive in the active site. The binding modes of ACC and the s tructural analog alanine specifically labeled with N-15 or O-17 were examin ed by using Q-band electron nuclear double resonance (ENDOR). The data indi cate that these molecules bind directly to the iron through both the alpha- amino and alpha-carboxylate groups. These observations are inconsistent wit h the currently favored mechanism for ACCO, in which it is proposed that bo th ascorbate and O-2 bind to the iron as a step in O-2 activation, We propo se a different mechanism in which the iron serves instead to simultaneously bind ACC and O-2, thereby fixing their relative orientations and promoting electron transfer between them to initiate catalysis.