作者: Thomas Wieland , Karl H Jakobs
DOI: 10.1016/S0076-6879(94)37048-6
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摘要: Publisher Summary This chapter discusses measurement of receptor-stimulated guanosine 5'-O-(γ-Thio)triphosphate binding by G Proteins. Many transmembrane signaling processes caused extracellular hormones and neurotransmitters are mediated receptors interacting with heterotrimeric (αβγ) guanine nucleotide-binding proteins (G proteins) attached to the inner face plasma membrane. Agonist-liganded apparently initiate activation catalyzing exchange 5'-diphosphate (GDP) 5'-triphosphate (GTP) bound α subunits. In membrane preparations reconstituted systems, this process is frequently monitored studying agonist stimulation high-affinity GTPase, an enzymatic activity G-protein To study initial steps agonist-liganded in a quantitative manner, radiolabeled GTP analogs, which not hydrolyzed GTPase subunits, determined. Of these 5'-O-(γ-[ 35 S]thio)triphosphate ([ S]GTPγS) most used. nucleotide has high affinity for all types available relatively specific radioactivity (1000-1400 Ci/mmol; physical half-life 87.4 days). The describe receptor induced [ S]GTPγS membranous detergent-solubilized how method can be adapted different optimal response stimulation.