Expression and activity of gamma‐glutamyl transpeptidase in the rat epididymis

Abstract

Following Northern analysis, GGT mRNA was found predominantly within the caput epididymides and kidney. The size of mRNAs for kidney, caput, corpus, and ductus 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 greater than cauda and corpus greater than caput greater than 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 microM and 2.19 nmoles/min/microliters luminal fluid at pH 8.5 compared to 0.49 microM and 0.49 nmoles/min/microliters 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 microM) and higher Vmax (1.13 nmoles/min/microliters 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.(ABSTRACT TRUNCATED AT 250 WORDS)

Contribution of epididymal secretory proteins for spermatozoa maturation

“…Glutathion (GSH) and oxidized glutathion (GSSG) are present in the epididymis (Hinton et al, 1996), as are glutathion-metabolizing enzymes such as glutathion peroxidase and gamma-glutamyl transpeptidase (GGT). These two enzymes are secreted in the proximal region, where their concentrations in the luminal fluid are the most abundant (DeLap et al, 1977;Hales et al, 1980;Kohdaira et al, 1986;Agrawal and Vanha-Perttula, 1988;Hinton et al, 1991), GPX is also associated with the sperm membrane (Jimenez et al, 1990;Perry et al, 1992).…”

Multiple Forms of Gamma-Glutamyl Transpeptidase Messenger Ribonucleic Acid are Expressed in the Adult Rat Testis and Epididymis1

“…The cleavage of glutathione by GGT and the subsequent hydrolysis of cys-gly by dipeptidase enable cells to reabsorb these amino acids, thus supporting the intracellular synthesis and subsequent export of reduced glutathione [19]. Epididymal GGT has a high affinity for oxidized glutathione and is likely to play a critical role in controlling the oxidized/reduced glutathione balance to maintain concentrations of reduced glutathione capable of performing antioxidant functions within the luminal fluid [20].…”

“…GGT is a heterodimeric integral membrane glycoprotein present on the outer surface of many epithelial cells, including epididymal epithelial cells, and within epididymal luminal fluid [19][20][21][22][23].…”

“…In the adult rat epididymis, GGT catalytic activity and histochemical localization markedly decrease from the initial segment to the cauda epididymidis [20][21][22][23][24][25]. To understand the cellular processes responsible for the regulation of GGT catalytic activity along the epididymal duct requires a knowledge of the molecular events that govern GGT mRNA expression in different regions of the epididymis.…”

Proteomic Profiling of Regionalized Proteins in Rat Epididymis Indicates Consistency between Specialized Distribution and Protein Functions

“…Several aspects of our results indicate that this exclusion was effective. For example, although γ-glutamyltransferase is in the luminal fluid throughout the epididymis, 26 it was detected only in the caput ( Figure 3A), which suggested that γ-glutamyltransferase in the luminal fluid had been removed. In addition, the results from our Northern blot analysis showed that nearly all of the selected proteins were expressed in situ.…”