Tamir H, Fawzi A B, Tamir A, Evans T, Northup J K
Department of Pharmacology, Yale University School of Medicine, New Haven, Connecticut 06510.
Biochemistry. 1991 Apr 23;30(16):3929-36. doi: 10.1021/bi00230a018.
Signal-transducing G-proteins are heterotrimers composed of GTP-binding alpha subunits in association with a tightly bound complex of beta and gamma subunits. While the alpha subunits are recognized as a family of diverse structures, beta and gamma subunits have also been found as heterogeneous isoforms. To investigate the diversity and tissue specificity of the beta gamma complexes, we have examined homogeneous oligomeric G-proteins from a variety of sources. The beta and gamma subunits isolated from the major-abundance G-proteins from bovine brain, bovine retina, rabbit liver, human placenta, and human platelets were purified and subjected to biochemical and immunological analysis. Protease mapping and immune recognition revealed an identical profile for each of the two distinctly migrating beta isoforms (beta 36 and beta 35) regardless of tissue or G-protein origin. Digestion with V8 protease revealed four distinct, clearly resolved terminal fragments for beta 36 and two for beta 35. Trypsin and chymotrypsin digestion yielded numerous bands, but again each form had a unique profile with no tissue specificity. Tryptic digestion was found to be conformationally specific with the most resistant structure being the native beta gamma complex. With increasing trypsin, the complex was digested but in a pattern distinct from that for denatured beta. In contrast to the two highly homologous beta structures, examination of this set of proteins revealed at least six distinct gamma peptides. Two unique gamma peptides were found in bovine retinal Gt and three gamma peptides in samples of bovine brain derived Go/Gi. Human placental and platelet Gi samples each contained a unique gamma.(ABSTRACT TRUNCATED AT 250 WORDS)
信号转导G蛋白是由GTP结合α亚基与紧密结合的β和γ亚基复合物组成的异源三聚体。虽然α亚基被认为是一个具有多种结构的家族，但β和γ亚基也被发现是异质同工型。为了研究βγ复合物的多样性和组织特异性，我们检测了来自多种来源的均一寡聚G蛋白。从牛脑、牛视网膜、兔肝、人胎盘和人血小板中主要丰度的G蛋白中分离出的β和γ亚基被纯化，并进行了生化和免疫学分析。蛋白酶图谱分析和免疫识别显示，两种明显迁移的β同工型（β36和β35）中的每一种都具有相同的图谱，而与组织或G蛋白来源无关。用V8蛋白酶消化显示β36有四个不同的、清晰分辨的末端片段，β35有两个。胰蛋白酶和胰凝乳蛋白酶消化产生了许多条带，但同样每种形式都有独特的图谱，没有组织特异性。发现胰蛋白酶消化具有构象特异性，最具抗性的结构是天然βγ复合物。随着胰蛋白酶用量增加，复合物被消化，但消化模式与变性β不同。与两种高度同源的β结构不同，对这组蛋白质的检测显示至少有六种不同的γ肽。在牛视网膜Gt中发现了两种独特的γ肽，在牛脑来源的Go/Gi样品中有三种γ肽。人胎盘和血小板Gi样品各自含有一种独特的γ肽。（摘要截短于250字）
