Expression and functional characterization of the cancer-related serine protease, human tissue kallikrein 14
Expression and functional characterization of the cancer-related serine protease, human tissue kallikrein 14
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DOI：
10.1074/jbc.m608348200
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发表时间：
2007-01-26
影响因子：
4.8
通讯作者：
Diamandis, Eleftherios P.
中科院分区：
文献类型：
--
作者：
Borgono, Carla A.;Michael, Iacovos P.;Diamandis, Eleftherios P.
人组织激肽释放酶14（KLK 14）是一种新的胞外丝氨酸蛋白酶。临床数据将KLK 14表达与几种疾病联系起来，主要是癌症;然而，对其（病理）生理作用知之甚少。为了表征KLK 14的功能，我们表达并纯化了成熟和酶原形式的重组KLK 14，并确定了其表达模式、特异性、调节和体外底物。通过使用我们的新型免疫测定，正常和/或患病的皮肤、乳房、前列腺和卵巢含有最高浓度的KLK 14。与健康男性相比，前列腺癌患者的血清KLK 14水平显著升高。KLK 14对P1-Arg的选择性高于对Lys的选择性，具有胰蛋白酶样的特异性。KLK 14活性的调节方式为：1）自溶裂解导致酶失活; 2）通过抑制性丝氨酸蛋白酶抑制剂α（1）-抗胰蛋白酶、α（2）-抗纤溶酶、抗凝血酶III和α 1-抗胰凝乳蛋白酶，二级速率常数（k（+2）/K-i）分别为49.8、23.8、1.48和0.224 μ M-1 min（-1），以及纤溶酶原激活物抑制剂-1; 3）柠檬酸盐和锌离子，其分别对KLK 14活性产生刺激和抑制作用。我们还扩展了KLK 14的体外靶谱，包括胶原蛋白I-IV、纤连蛋白、层粘连蛋白、激肽原、纤维蛋白原、纤溶酶原、玻连蛋白和胰岛素样生长因子结合蛋白2和3。我们的研究结果表明，KLK 14可能与肿瘤进展的几个方面有关，包括生长，侵袭和血管生成，以及通过软骨退化参与关节炎疾病。这些发现可能对癌症和其他KLK 14活性升高的疾病的管理具有临床意义。
Human tissue kallikrein 14 (KLK14) is a novel extracellular serine protease. Clinical data link KLK14 expression to several diseases, primarily cancer; however, little is known of its (patho)-physiological role. To functionally characterize KLK14, we expressed and purified recombinant KLK14 in mature and proenzyme forms and determined its expression pattern, specificity, regulation, and in vitro substrates. By using our novel immunoassay, the normal and/or diseased skin, breast, prostate, and ovary contained the highest concentration of KLK14. Serum KLK14 levels were significantly elevated in prostate cancer patients compared with healthy males. KLK14 displayed trypsin-like specificity with high selectivity for P1-Arg over Lys. KLK14 activity could be regulated as follows: 1) by autolytic cleavage leading to enzymatic inactivation; 2) by the inhibitory serpins alpha(1)-antitrypsin, alpha(2)-antiplasmin, antithrombin III, and alpha 1-antichymotrypsin with second order rate constants (k(+2)/K-i) of 49.8, 23.8, 1.48, and 0.224 mu M-1 min(-1), respectively, as well as plasminogen activator inhibitor-1; and 3) by citrate and zinc ions, which exerted stimulatory and inhibitory effects on KLK14 activity, respectively. We also expanded the in vitro target repertoire of KLK14 to include collagens I-IV, fibronectin, laminin, kininogen, fibrinogen, plasminogen, vitronectin, and insulin-like growth factor-binding proteins 2 and 3. Our results indicate that KLK14 may be implicated in several facets of tumor progression, including growth, invasion, and angiogenesis, as well as in arthritic disease via deterioration of cartilage. These findings may have clinical implications for the management of cancer and other disorders in which KLK14 activity is elevated.