Nishi N, Tanabe H, Oya H, Urushihara M, Miyanaka H, Wada F
Department of Endocrinology, Kagawa Medical School, Japan.
J Biol Chem. 1994 Jan 14;269(2):1015-9.
We reported previously that a monoclonal antibody against probasin (rat prostatic secretory protein) recognizes a 40-kDa protein localized in rat liver and kidney. The protein (probasin-related antigen, PRB-RA) may participate in a specific differentiated function of these tissues. To clarify the molecular nature of PRB-RA, a series of cDNA clones coding for the protein were isolated from a rat liver expression library using an affinity-purified polyclonal antibody. The amino acid sequence deduced from the determined cDNA sequence included sequences identical with those of proteolytic fragments of PRB-RA, which covered about 70% of the deduced sequence. Northern blot hybridization of poly(A)+ RNA isolated from rat tissues showed the presence of predominant and minor mRNA species of about 2.0 and 4.3 kilobases, respectively, in the liver and kidney. A sequence homology search revealed that PRB-RA is almost completely identical to rat cystathionine gamma-lyase (cystathionase) and that it does not show overall homology with probasin. Three candidates for an epitope common to probasin and PRB-RA were found on close examination of the amino acid sequences of the two proteins. A synthetic peptide, TYFRRI, corresponding to one of the candidates, neutralized the reactivity of the anti-probasin monoclonal antibody to both probasin and PRB-RA on Western blot analysis. These results show that PRB-RA/cystathionase is neither structurally nor functionally related to probasin except for a common epitope and that cystathionase, a cystein-producing enzyme, is localized in urinary tubular epithelial cells in a highly restricted region of the kidney in addition to in liver parenchymal cells.
我们先前报道过，一种抗前列腺素（大鼠前列腺分泌蛋白）的单克隆抗体可识别一种定位于大鼠肝脏和肾脏的40 kDa蛋白。该蛋白（前列腺素相关抗原，PRB - RA）可能参与这些组织的特定分化功能。为阐明PRB - RA的分子本质，使用亲和纯化的多克隆抗体从大鼠肝脏表达文库中分离出一系列编码该蛋白的cDNA克隆。从所确定的cDNA序列推导的氨基酸序列包含与PRB - RA蛋白水解片段相同的序列，这些片段覆盖了推导序列的约70%。从大鼠组织分离的poly(A)+ RNA的Northern印迹杂交显示，在肝脏和肾脏中分别存在约2.0和4.3千碱基的主要和次要mRNA种类。序列同源性搜索表明，PRB - RA与大鼠胱硫醚γ - 裂解酶（胱硫醚酶）几乎完全相同，且与前列腺素没有整体同源性。仔细检查这两种蛋白的氨基酸序列后，发现了前列腺素和PRB - RA共有的三个潜在表位。在蛋白质印迹分析中，对应其中一个潜在表位的合成肽TYFRRI可中和抗前列腺素单克隆抗体对前列腺素和PRB - RA的反应性。这些结果表明，除了一个共同表位外，PRB - RA/胱硫醚酶在结构和功能上与前列腺素均无关联，并且胱硫醚酶这种产生半胱氨酸的酶，除了存在于肝实质细胞中外，还高度局限地定位于肾脏的肾小管上皮细胞中。
