Molecular organization of bovine rod cGMP-phosphodiesterase 6

Phosphodiesterase 6 (PDE6), a multisubunit (alpha beta gamma (2)delta) enzy me, plays a major role in visual function by hydrolysing cGMP in response t o a light stimulus. Solubilized bovine rod PDE6 molecules depleted of their gamma subunits were purified to homogeneity from bovine retinal rods and t heir molecular organization was investigated by electron microscopy. Image analysis of single particles revealed the three-dimensional dimeric arrange ment of the purified alpha beta delta complex, and the internal organizatio n of each catalytic subunit into three distinct domains at a resolution of 2.8 nm. The relative volume of each domain is consistent with sequence anal ysis and functional data, which suggest that these domains correspond to th e catalytic and two CAF domains. This hypothesis was confirmed by immunolab elling experiments, which located the N-terminal part of the catalytic subu nit where the major interaction between the two alpha beta subunits was fou nd to occur. The 3D molecular organization of human platelet PDE5 appears h ighly homologous to that of bovine rod PDE6, as predicted by similarities i n their primary sequences. These observations describe the quaternary organ ization of the catalytic PDE6 ccp complex, and place the catalytic and regu latory domains on a structural model. (C) 2001 Academic Press.