NMR Structure of Free RGS4 Reveals an Induced Conformational Change upon Binding G

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• 作者：Franklin J. Moy ; Pranab K. Chanda ; Mark I. Cockett ; Wade Edris ; Philip G. Jones ; Kim Mason ; Simon Semus ; and Robert Powers
• 刊名：Biochemistry
• 出版年：2000
• 出版时间：June 20, 2000
• 年：2000
• 卷：39
• 期：24
• 页码：7063 - 7073
• 全文大小：434K
• 年卷期：v.39,no.24(June 20, 2000)
• ISSN：1520-4995

文摘

Heterotrimeric guanine nucleotide-binding proteins (G-proteins) are transducers in many cellulartransmembrane signaling systems where regulators of G-protein signaling (RGS) act as attenuators of theG-protein signal cascade by binding to the Gimages/gifchars/alpha.gif" BORDER=0> subunit of G-proteins (G_{iimages/gifchars/alpha.gif" BORDER=0>1}) and increasing the rate ofGTP hydrolysis. The high-resolution solution structure of free RGS4 has been determined using two-dimensional and three-dimensional heteronuclear NMR spectroscopy. A total of 30 structures werecalculated by means of hybrid distance geometry-simulated annealing using a total of 2871 experimentalNMR restraints. The atomic rms distribution about the mean coordinate positions for residues 5-134 forthe 30 structures is 0.47 ± 0.05 Å for the backbone atoms, 0.86 ± 0.05 Å for all atoms, and 0.56 ± 0.04Å for all atoms excluding disordered side chains. The NMR solution structure of free RGS4 suggests asignificant conformational change upon binding G_{iimages/gifchars/alpha.gif" BORDER=0>1} as evident by the backbone atomic rms differenceof 1.94 Å between the free and bound forms of RGS4. The underlying cause of this structural change isa perturbation in the secondary structure elements in the vicinity of the G_{iimages/gifchars/alpha.gif" BORDER=0>1} binding site. A kink in thehelix between residues K116-Y119 is more pronounced in the RGS4-G_{iimages/gifchars/alpha.gif" BORDER=0>1} X-ray structure relative tothe free RGS4 NMR structure, resulting in a reorganization of the packing of the N-terminal and C-terminalhelices. The presence of the helical disruption in the RGS4-G_{iimages/gifchars/alpha.gif" BORDER=0>1} X-ray structure allows for the formationof a hydrogen-bonding network within the binding pocket for G_{iimages/gifchars/alpha.gif" BORDER=0>1} on RGS4, where RGS4 residues D117,S118, and R121 interact with residue T182 from G_{iimages/gifchars/alpha.gif" BORDER=0>1}. The binding pocket for G_{iimages/gifchars/alpha.gif" BORDER=0>1} on RGS4 is larger andmore accessible in the free RGS4 NMR structure and does not present the preformed binding site observedin the RGS4-G_{iimages/gifchars/alpha.gif" BORDER=0>1} X-ray structure. This observation implies that the successful complex formation betweenRGS4 and G_{iimages/gifchars/alpha.gif" BORDER=0>1} is dependent on both the formation of the bound RGS4 conformation and the properorientation of T182 from G_{iimages/gifchars/alpha.gif" BORDER=0>1}. The observed changes for the free RGS4 NMR structure suggest a mechanismfor its selectivity for the Gimages/gifchars/alpha.gif" BORDER=0>-GTP-Mg²⁺ complex and a means to facilitate the GTPase cycle.