Yan K, Gautam N
Department of, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
J Biol Chem. 1997 Jan 24;272(4):2056-9. doi: 10.1074/jbc.272.4.2056.
In the yeast two-hybrid system, a 100-residue fragment (beta1A) from the N terminus of the beta1 subunit interacts with domains specific to adenylyl cyclase 2 (AC2), the muscarinic atrial potassium channel (GIRK1), and phospholipase C-beta2 (PLC-beta2). Based on the crystal structure of the G protein, beta1A is composed of an N-terminal alpha helix, a loop, and five beta strands in which the C-terminal four beta strands form a beta sheet, the first of seven sheets that make up the propeller structure of the beta subunit. A mutant of beta1A (L4P, L7P, and L14P), in which the alpha helix was potentially destroyed, interacted poorly with the G protein gamma subunit but effectively with domains of AC2, GIRK1, and PLC-beta2. In contrast, another mutant of beta1A (S72A, D76A, and W82A), in which a network of hydrogen bonds was disrupted, interacted poorly with GIRK1 and PLC-beta2 domains, but effectively with the gamma subunit and the AC2 domain. These results suggest that the proper folding of the first five beta strands in the G protein beta subunit is a requirement for appropriately positioning residues that interact with GIRK1 and PLC-beta2. Furthermore, since mutations that potentially disrupted the folding of these beta strands did not affect interaction with AC2, the structural determinants on the G protein beta subunit for interaction with various effectors may be different.
在酵母双杂交系统中，β1亚基N端的一个100个氨基酸残基的片段（β1A）与腺苷酸环化酶2（AC2）、毒蕈碱型心房钾通道（GIRK1）和磷脂酶C-β2（PLC-β2）的特定结构域相互作用。基于G蛋白的晶体结构，β1A由一个N端α螺旋、一个环和五条β链组成，其中C端的四条β链形成一个β折叠片层，这是构成β亚基螺旋桨结构的七个片层中的第一个。β1A的一个突变体（L4P、L7P和L14P），其中α螺旋可能被破坏，与G蛋白γ亚基的相互作用较差，但与AC2、GIRK1和PLC-β2的结构域有效相互作用。相比之下，β1A的另一个突变体（S72A、D76A和W82A），其中氢键网络被破坏，与GIRK1和PLC-β2结构域的相互作用较差，但与γ亚基和AC2结构域有效相互作用。这些结果表明，G蛋白β亚基中前五条β链的正确折叠是与GIRK1和PLC-β2相互作用的残基正确定位的必要条件。此外，由于可能破坏这些β链折叠的突变并不影响与AC2的相互作用，G蛋白β亚基与各种效应器相互作用的结构决定因素可能不同。
