Brömme D, Rossi A B, Smeekens S P, Anderson D C, Payan D G
Khepri Pharmaceuticals, Inc., South San Francisco, California 94080, USA.
Biochemistry. 1996 May 28;35(21):6706-14. doi: 10.1021/bi960092y.
We have cloned the cDNA of human bleomycin hydrolase (hBH), a protease which is thought to be involved in the metabolic inactivation of the antineoplastic drug bleomycin. The open reading frame consists of 1365 base pairs and is predicted to encode a 52 kDa protein. The protein shares 40% identity with yeast bleomycin hydrolase and contains the conserved active site residues (Cys, His, Asn) characteristic for cysteine proteases of the papain superfamily. Human bleomycin hydrolase has been functionally expressed in Spodoptera frugiperda Sf9 cells using the Autographa californica nuclear polyhedrosis virus. The 52 kDa recombinant protein forms a hexamer of 310 kDa and acts strictly as an aminopeptidase with a broad substrate specificity. The lack of a leader sequence and its pH optimum at 7.2 suggest a cytosolic/nuclear localization. Human bleomycin hydrolase was detected at low to moderate expression levels in most of the human organs tested. Significantly higher RNA levels have been observed in a variety of tumor cell lines. The human enzyme effectively degrades both forms of bleomycin (A2 and B2) in vitro and could indeed be responsible for the resistance of various tumors to this widely used anticancer drug.
我们已经克隆了人博来霉素水解酶（hBH）的cDNA，这是一种蛋白酶，被认为参与了抗肿瘤药物博来霉素的代谢失活过程。其开放阅读框由1365个碱基对组成，预计编码一个52 kDa的蛋白质。该蛋白质与酵母博来霉素水解酶有40%的同源性，并含有木瓜蛋白酶超家族半胱氨酸蛋白酶特有的保守活性位点残基（半胱氨酸、组氨酸、天冬酰胺）。人博来霉素水解酶已利用苜蓿银纹夜蛾核型多角体病毒在草地贪夜蛾Sf9细胞中实现功能表达。这种52 kDa的重组蛋白形成一个310 kDa的六聚体，并且严格作为一种具有广泛底物特异性的氨肽酶发挥作用。缺乏前导序列以及其最适pH为7.2表明其定位于胞质/细胞核。在大多数检测的人体器官中，人博来霉素水解酶以低至中等水平表达。在多种肿瘤细胞系中观察到其RNA水平显著更高。该人源酶在体外能有效降解两种形式的博来霉素（A2和B2），确实可能是各种肿瘤对这种广泛使用的抗癌药物产生耐药性的原因。
