PMID- 18467334 OWN - NLM STAT- MEDLINE DCOM- 20080828 LR - 20231213 IS - 0021-9258 (Print) IS - 1083-351X (Electronic) IS - 0021-9258 (Linking) VI - 283 IP - 28 DP - 2008 Jul 11 TI - Juxtaposition of the steroid binding domain-like I and II regions constitutes a ligand binding site in the sigma-1 receptor. PG - 19646-56 LID - 10.1074/jbc.M802192200 [doi] AB - sigma-1 receptors represent unique binding sites that are capable of interacting with a wide range of compounds to mediate different cellular events. The composition of the ligand binding site of this receptor is unclear, since no NMR or crystal structures are available. Recent studies in our laboratory using radiolabeled photoreactive ligands suggested that the steroid binding domain-like I (SBDLI) (amino acids 91-109) and the steroid binding domain-like II (SBDLII) (amino acids 176-194) regions are involved in forming the ligand binding site(s) ( Chen, Y., Hajipour, A. R., Sievert, M. K., Arbabian, M., and Ruoho, A. E. (2007) Biochemistry 46, 3532-3542 ; Pal, A., Hajipour, A. R., Fontanilla, D., Ramachandran, S., Chu, U. B., Mavlyutov, T., and Ruoho, A. E. (2007) Mol. Pharmacol. 72, 921-933 ). In this report, we have further addressed this issue by utilizing our previously developed sulfhydryl-reactive, cleavable, radioiodinated photocross-linking reagent: methanesulfonothioic acid, S-((4-(4-amino-3-[125I]iodobenzoyl) phenyl)methyl) ester (Guo, L. W., Hajipour, A. R., Gavala, M. L., Arbabian, M., Martemyanov, K. A., Arshavsky, V. Y., and Ruoho, A. E. (2005) Bioconjugate Chem. 16, 685-693). This photoprobe was shown to derivatize the single cysteine residues as mixed disulfides at position 94 in the SBDLI region of the wild type guinea pig sigma-1 receptor (Cys94) and at position 190 in the SBDLII region of a mutant guinea pig sigma-1 receptor (C94A,V190C), both in a sigma-ligand (haloperidol or (+)-pentazocine)-sensitive manner. Significantly, photocross-linking followed by Endo Lys-C cleavage under reducing conditions and intramolecular radiolabel transfer from the SBDLI to the SBDLII region in the wild type receptor and, conversely, from the SBDLII to the SBDLI region in the mutant receptor were observed. These data support a model in which the SBDLI and SBDLII regions are juxtaposed to form, at least in part, a ligand binding site of the sigma-1 receptor. FAU - Pal, Arindam AU - Pal A AD - Department of Pharmacology, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin 53705, USA. FAU - Chu, Uyen B AU - Chu UB FAU - Ramachandran, Subramaniam AU - Ramachandran S FAU - Grawoig, David AU - Grawoig D FAU - Guo, Lian-Wang AU - Guo LW FAU - Hajipour, Abdol R AU - Hajipour AR FAU - Ruoho, Arnold E AU - Ruoho AE LA - eng GR - R01 MH065503/MH/NIMH NIH HHS/United States PT - Journal Article PT - Research Support, N.I.H., Extramural DEP - 20080507 PL - United States TA - J Biol Chem JT - The Journal of biological chemistry JID - 2985121R RN - 0 (Disulfides) RN - 0 (Ligands) RN - 0 (Receptors, sigma) RN - 0 (Recombinant Proteins) RN - 0 (Steroids) RN - K848JZ4886 (Cysteine) SB - IM MH - Animals MH - Cysteine/chemistry/genetics/metabolism MH - Disulfides/chemistry/metabolism MH - Guinea Pigs MH - Ligands MH - *Models, Molecular MH - Protein Binding/genetics MH - Protein Structure, Tertiary/genetics MH - Receptors, sigma/*chemistry/genetics/metabolism MH - Recombinant Proteins/chemistry/genetics/metabolism MH - Steroids/*chemistry/metabolism MH - Sigma-1 Receptor PMC - PMC2443669 EDAT- 2008/05/10 09:00 MHDA- 2008/08/30 09:00 PMCR- 2009/07/11 CRDT- 2008/05/10 09:00 PHST- 2008/05/10 09:00 [pubmed] PHST- 2008/08/30 09:00 [medline] PHST- 2008/05/10 09:00 [entrez] PHST- 2009/07/11 00:00 [pmc-release] AID - S0021-9258(20)81418-5 [pii] AID - 19646 [pii] AID - 10.1074/jbc.M802192200 [doi] PST - ppublish SO - J Biol Chem. 2008 Jul 11;283(28):19646-56. doi: 10.1074/jbc.M802192200. Epub 2008 May 7.