A carboxypeptidase inhibitor from the tick Rhipicephalus bursa
A carboxypeptidase inhibitor from the tick Rhipicephalus bursa
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DOI：
10.1074/jbc.m411086200
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发表时间：
2005-02-04
影响因子：
4.8
通讯作者：
Sommerhoff, CP
中科院分区：
文献类型：
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作者：
Arolas, JL;Lorenzo, J;Sommerhoff, CP
从囊扇头蜱（Rhipicephalus bursa）中分离得到一种新的金属羧肽酶抑制剂，命名为蜱羧肽酶抑制剂（TCI）。通过逆转录-PCR和cDNA末端快速扩增技术从蜱mRNA中克隆了编码该蛋白的完整cDNA。全长TCI cDNA含有编码97个氨基酸残基的前体蛋白的开放阅读框架，所述前体蛋白由22个残基的预测信号肽和成熟TCI（75个残基的富含半胱氨酸的蛋白质（12个Cys））组成。推导的氨基酸序列显示没有同源性，其他已知的蛋白质; C末端，但是，类似于那些其他蛋白质金属羧肽酶抑制剂，这表明一个共同的抑制机制。在大肠杆菌中表达的重组TCI是完全功能性的，并且抑制A/B亚家族的羧肽酶，其平衡解离常数在纳摩尔范围内。圆二色谱和核磁共振结构分析。共振表明TCI是一种受二硫键强烈约束的蛋白质，在宽pH范围内异常稳定，并且对变性条件具有高度抗性。作为血浆羧肽酶B的紧密结合抑制剂，也称为凝血酶激活的纤维蛋白溶解抑制剂，重组TCI在体外刺激纤维蛋白溶解，因此可能具有应用于预防或治疗血栓性疾病的潜力。
A novel proteinaceous metallo-carboxypeptidase inhibitor, named tick carboxypeptidase inhibitor (TCI), was isolated from the ixodid tick Rhipicephalus bursa and N-terminally sequenced. The complete cDNA encoding this protein was cloned from tick mRNA by reverse transcription-PCR and rapid amplification of cDNA ends techniques. The full-length TCI cDNA contains an open reading frame coding for a precursor protein of 97 amino acid residues that consists of a predicted signal peptide of 22 residues and of mature TCI, a 75-residue cysteine-rich protein (12 Cys). The deduced amino acid sequence shows no homology to other known proteins; the C terminus, however, resembles those of other protein metallo-carboxypeptidase inhibitors, suggesting a common mechanism of inhibition. Recombinant TCI expressed in Escherichia coli is fully functional and inhibits carboxypeptidases of the A/B subfamily with equilibrium dissociation constants in the nanomolar range. Structural analyses by circular dichroism and nuclear magnetic. resonance indicate that TCI is a protein strongly constrained by disulfide bonds, unusually stable over a wide pH range and highly resistant to denaturing conditions. As a tight binding inhibitor of plasma carboxypeptidase B, also known as thrombin-activatable fibrinolysis inhibitor, recombinant TCI stimulates fibrinolysis in vitro and thus may have potential for applications to prevent or treat thrombotic disorders.