Use of 5'-[p-(fluorosulfonyl)benzoyl] guanosine as an affinity probe for the guanine nucleotide-binding site of transducin.
Transducin (T) mediates vision in retinal rods by transmitting light signals detected by rhodopsin to a cGMP phosphodiesterase. The flow of information relies on a subunit association/dissociation cycle of T regulated by a guanine nucleotide exchange/hydrolysis reaction. 5'-[p-(Fluorosulfonyl)benzoyl] guanosine (FSBG) was synthesized and examined here as an affinity label for the guanine nucleotide binding site of T. Although the relative binding affinity of FSBG to T was much lower than for GTP and beta,gamma-imido-guanosine 5'-triphosphate (GMPPNP), the incorporation of FSBG to T inhibited its light-dependent [(3)H] GMPPNP binding activity in a concentration dependent manner. Additionally, GDP, GTP and GTP analogs hindered the binding of [(3)H] FSBG to T. These results demonstrated that FSBG could be used to specifically modify the active site of T. In addition, FSBG was not capable of dissociating T from T:photoactivated rhodopsin complexes, suggesting that in this case FSBG is acting as a GDP analog.
Jaffé
Matthias
M
Departamento de Química, Universidad Simón Bolívar, Valle de Sartenejas, Caracas, Venezuela.
Bubis
José
J
D000345:Affinity Labels; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D006147:Guanine / Q000737:chemistry / Q000378:metabolism*; D006151:Guanosine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D015293:Transducin / Q000737:chemistry* / Q000378:metabolism*
C017320:5'-(4-fluorosulfonylbenzoyl)guanosine; D000345:Affinity Labels; D006151:Guanosine; D006147:Guanine; D015293:Transducin