Expression and activity of gamma-glutamyl transpeptidase in the rat epididymis.
Expression and activity of gamma-glutamyl transpeptidase in the rat epididymis.
复制标题
大鼠附睾中γ-谷氨酰转肽酶的表达和活性。
DOI：
10.1002/mrd.1080280107
复制
发表时间：
1991
影响因子：
2.5
通讯作者：
Good,K
中科院分区：
文献类型：
--
作者：
Hinton,BT;Palladino,MA;Mattmueller,DR;Bard,D;Good,K
Northern分析显示，GGT mRNA主要存在于附睾头和肾脏中。肾、头、体、管mrna的大小分别为2.2、2.3、2.2。尾部则分别为2.2和2.3 kb。微穿刺采集附睾管腔液GGT转肽化和水解活性分别显示：caput=体>尾和体>尾和体>尾。丝氨酸‐硼酸盐显著抑制了GGT转肽酶活性，并在pH 8.0时达到最佳。在pH为8.5时，计算得到的GSH水解Km和Vmax分别为0.06 μM和2.19 nmol /min/μl，而在生理pH为6.5时，计算得到的GSH水解Km和Vmax分别为0.49 μM和0.49 nmol /min/μl。实验结果表明，GSSG与GSH相比，可降低GSSG的Km值（0.12 μM）和Vmax值（1.13 nmol /min/μl）。Triton X‐114分配实验的结果表明，管腔GGT可能以膜结合和非膜结合形式存在。Western blot分析附睾腔液中的蛋白，发现所有附睾区域都存在GGT亚基。然而，在尾液中也观察到两个较低的分子带，约为22 kDa和21 kDa。这表明，当GGT沿着附睾管运输时，它会发生降解，这是它在附睾远端区域失去活性的原因。附睾GGT可能不参与L -谷氨酸的运输，因为运输与沿附睾管的GGT mRNA表达程度无关。
Following Northern analysis, GGT mRNA was found predominantly within the caput epididymides and kidney. The size of mRNAs for kidney, caput, corpus, and ducts deferens were 2.2, 2.3, 2.2., and 2.3 kb, respectively, whereas cauda showed a doublet of 2.2 and 2.3 kb. GGT transpeptidation and hydrolytic activity within epididymal luminal fluids collected by micropuncture showed caput=corpus>cauda and corpus>caput>cauda, respectively. Caput luminal GGT transpeptidation activity was significantly inhibited by serine‐borate and was optimal at pH 8.0. The calculated Km and Vmax values for hydrolysis of GSH by caput luminal GGT were 0.06 μM and 2.19 nmoles/min/μl luminal fluid at pH 8.5 compared to 0.49 μM and 0.49 nmoles/min/μl luminal fluid, respectively, at the physiological pH 6.5 of caput fluid. These studies would suggest that the epididymis can control the activity of luminal GGT by pH. Lower Km (0.12 μM) and higher Vmax (1.13 nmoles/min/μl luminal fluid) values were also calculated when GSSG was used compared to GSH. Results from Triton X‐114 partitioning experiments suggest that luminal GGT probably exists in both membrane bound and nonmembrane bound forms. Western blot analysis of proteins within epididymal luminal fluids revealed both subunits of GGT in all epididymal regions studied. However, two lower molecular bands, approximately 22 kDa and 21 kDa, were also observed in cauda fluid. It is suggested that as GGT is transported along the epididymal duct it undergoes degradation, which accounts for its loss of activity in the distal epididymal regions. Epididymal GGT may not be involved in the transport of L‐glutamate since transport was not related to the degree of GGT mRNA expression along the epididymal duct.