Warsop H, Khouja A, Wichelhaus D P, Jones C T
University of Oxford, Laboratory of Cellular and Developmental Physiology, UK.
J Dev Physiol. 1993 Mar;19(3):91-7.
G-protein content (G(i) alpha, Gs alpha, Gq/11 alpha G(o) alpha and beta subunits) has been measured in membranes prepared from guinea pig uterus at different stages of pregnancy using SDS-PAGE and immunoblotting. Quantification using HRP- or 125I-labelled IgG as second antibody showed a good correlation between added membrane protein and measured G-protein content. Gs alpha appears as two bands of 45 kDa and 52 kDa respectively, the content of both were comparatively high in the non-pregnant uterus and fell about 4-fold close to term (60-67 days). G(i) alpha showed the converse with low level in membranes from the non-pregnant uterus with level approximately 6-fold higher by term. G(o) alpha exhibited changes similar to G(i) alpha. The changes in the content of Gq/11 alpha where biphasic, with comparatively high levels in membranes from the non-pregnant uterus, a sharp fall early in pregnancy followed by a 3-fold rise by near term. The uterine membrane content of the common beta subunit exhibited changes comparable to that of G(i) alpha and G(o) alpha with a 6-fold rise in content between non- and late-pregnant. Measurement of the effect of GTP gamma S action on phosphatidylinositol phospholipase C activity in uterine membranes with exogenous substrate showed pregnancy-dependent effects. In membranes from the non-pregnant uterus 0.1 microM GTP gamma S caused a modest stimulation of activity of 16 +/- 1.9%, whilst at 100 microM it inhibited the enzyme by 25 +/- 6.48%. In membranes from the late-pregnant guinea pig uterus GTP gamma S at both concentrations caused stimulation of enzyme activity.(ABSTRACT TRUNCATED AT 250 WORDS)
利用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）和免疫印迹法，测定了妊娠不同阶段豚鼠子宫制备的膜中G蛋白含量（G(i)α、Gsα、Gq/11α、G(o)α和β亚基）。使用辣根过氧化物酶（HRP）或125I标记的IgG作为二抗进行定量分析，结果显示添加的膜蛋白与测得的G蛋白含量之间具有良好的相关性。Gsα分别呈现为45 kDa和52 kDa的两条带，在未孕子宫中两者含量相对较高，接近足月时（60 - 67天）下降约4倍。G(i)α则相反，未孕子宫膜中的水平较低，足月时约高6倍。G(o)α呈现出与G(i)α相似的变化。Gq/11α含量的变化呈双相性，未孕子宫膜中的水平相对较高，妊娠早期急剧下降，随后在接近足月时上升3倍。共同β亚基的子宫膜含量变化与G(i)α和G(o)α相当，未孕和妊娠晚期之间含量上升6倍。用外源底物测定GTPγS作用对子宫膜中磷脂酰肌醇磷脂酶C活性的影响，结果显示出与妊娠相关的效应。在未孕子宫膜中，0.1 μM GTPγS引起活性适度刺激，为16±1.9%，而在100 μM时则抑制该酶活性25±6.48%。在妊娠晚期豚鼠子宫膜中，两种浓度的GTPγS均引起酶活性的刺激。（摘要截短于250字）
