Human HtrA, an evolutionarily conserved serine protease identified as a differentially expressed gene product in osteoarthritic cartilage
Human HtrA, an evolutionarily conserved serine protease identified as a differentially expressed gene product in osteoarthritic cartilage
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DOI：
10.1074/jbc.273.51.34406
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发表时间：
1998-12-18
影响因子：
4.8
通讯作者：
Crowl, RM
中科院分区：
文献类型：
--
作者：
Hu, SI;Carozza, M;Crowl, RM
通过对骨关节炎软骨中表达的转录本进行差异显示分析，鉴定了大肠杆菌htrA基因产物的人类同源物。该转录物先前被鉴定为在SV 40转化的成纤维细胞中被抑制（Zumbrunn，J，和Trueb，B，（1996）FEBS Lett. 398，187-192）。与非关节炎对照相比，骨关节炎个体软骨中HtrA mRNA水平升高约7倍，通过软骨提取物的免疫印迹分析证实了人HtrA蛋白的差异表达，在异源系统中表达的人HtrA蛋白被分泌并表现出内切蛋白水解活性，包括自催化切割。通过将推定的活性位点丝氨酸328突变成丙氨酸，消除了酶活性。还发现丝氨酸328是与α（1）-抗胰蛋白酶形成稳定复合物所必需的。我们已经确定HtrA基因在哺乳动物物种中是高度保守的：来自牛、兔和豚鼠的HtrA cDNA克隆编码的氨基酸序列与人的相同性为98%。在大肠在大肠杆菌中，热休克或氧化应激后细胞存活所需的功能性htrA基因产物;其作用似乎是降解变性蛋白。我们建议，哺乳动物HtrA，在进化过程中增加了一个新的功能，即mac 25同源结构域，在细胞生长调节中起着重要的作用。
The human homologue of the Escherichia coli htrA gene product was identified by the differential display analysis of transcripts expressed in osteoarthritic cartilage. This transcript was identified previously as being repressed in SV40-transformed fibroblasts (Zumbrunn, J,, and Trueb, B, (1996) FEBS Lett. 398, 187-192). Levels of HtrA mRNA were elevated similar to 7-fold in cartilage from individuals with osteoarthritis compared with nonarthritic controls, Differential expression of human HtrA protein was confirmed by an immunoblot analysis of cartilage extracts, Human HtrA protein expressed in heterologous systems was secreted and exhibited endoproteolytic activity, including autocatalytic cleavage. Conversion by mutagenesis of the putative active site serine 328 to alanine eliminated the enzymatic activity. Serine 328 was also found to be required for the formation of a stable complex with alpha(1)-antitrypsin. We have determined that the HtrA gene is highly conserved among mammalian species: the amino acid sequences encoded by HtrA cDNA clones from cow, rabbit, and guinea pig are 98% identical to human. In E. coli, a functional htrA gene product is required for cell survival after heat shock or oxidative stress; its role appears to be the degradation of denatured proteins. We propose that mammalian HtrA, with the addition of a new functionality during evolution, i.e. a mac25 homology domain, plays an important role in cell growth regulation.