30-41 LS1_main_02_new.qxp

Rhodopsin is the primary molecule in the visual signaling cascade in vertebrate and invertebrate photoreceptor cells. Upon light absorption, the retinal chromophore in rhodopsin undergoes isomerization from the 11-cis to the all-trans configuration, which initiates the photoactivation process. In vertebrate retinas, the final photoproduct, meta-rhodopsin, activates a subtype of heterotrimeric G-protein, transducin (Gt), which in turn activates phosphodiesterase, an enzyme that degrades the second messenger, cGMP. In the retinas of many invertebrates such as insects and cephalopods, on the other hand, photoactivated acid meta-rhodopsin stimulates a Gq-type G-protein, and then the Gprotein activates phospholipase C, an enzyme that hydrolyzes phosphatidyl inositol into the second messengers inositol trisphosphate and diacyl glycerol [1]. The same cascade is used by many G-proteincoupled hormone/neurotransmitter receptors such as serotonin and acetylcholine receptors, indicating that invertebrate rhodopsin is a prototypical member of the G-protein-coupled receptor (GPCR) family. The structural information of invertebrate rhodopsin would be useful to elucidate a common mechanism of Gq-type G-protein activation by GPCR. We performed X-ray crystallographic analysis of invertebrate squid (Todarodes pacificus) rhodopsin. Squid rhodopsin contains 448 amino acids, i.e., 100 residues longer than those of vertebrate counterparts, mainly due to the unique C-terminal extension with repeated proline-rich sequences. After enzymatic deletion of the extended C-terminus, squid rhodopsin was extracted selectively from microvillar membrane with octylglucoside in the presence of zinc ion, and crystallized into the hexagonal P62 crystal. We collected diffraction data from a frozen crystal of squid rhodopsin at beamline BL38B1 and determined its crystal structure at 2.5 Å resolution [2]. The polypeptide of C-terminally truncated squid rhodopsin, which was traced from Glu 9 to Glu 358, is composed of seven transmembrane helices (from I to VII) and two cytoplasmic helices (VIII and IX) (Fig. 1). The overall structure revealed new features of Gqcoupled receptors. Helices V and VI extend into the cytoplasmic medium and, together with a hydrophilic helix IX, they form a highly organized structure of about 25 Å length from the membrane surface. The calculated map of electrostatic potential on the protein