Sakuma T, Higashiyama S, Hosoe S, Hayashi S, Taniguchi N
Department of Biochemistry, Osaka University Medical School, Suita.
J Biochem. 1997 Aug;122(2):474-80. doi: 10.1093/oxfordjournals.jbchem.a021776.
Heparin/heparan-sulfate proteoglycan (HSPG) binds to heparin-binding epidermal growth factor-like growth factor (HB-EGF) through its heparin-binding domain (HBD), which consists of 21 amino acid residues (P21). The CD9 antigen also interacts with a membrane-anchored form of HB-EGF (proHB-EGF) and enhances its juxtacrine activity. The CD9 antigen potentiates the juxtacrine activity of both proHB-EGF and proamphiregulin, but has no effect on proTGF-alpha. While both HB-EGF and amphiregulin contain an HBD, TGF-alpha does not. This suggests that the HBD of HB-EGF is also involved in CD9 antigen binding. Mutant CHO cells which lack HSPG recovered their capacity to bind to immobilized P21 when transfected with CD9 antigen cDNA. This binding was competitively inhibited by heparin in a dose-dependent manner. The interactions between synthetic peptides corresponding to the extracellular domain of CD9 antigen and the immobilized P21 were analyzed with surface plasmon resonance. The 119VIKEVQEFYKDTYNKLKTKD138 sequence of the CD9 antigen is thought to represent the binding site for HB-EGF. The k(D) values for heparin/P21 and 119V-D138/P21 were (2.82+/-0.10) x 10(-8) M and (3.71+/-0.71) x 10(-5) M, respectively. These results suggest that the 119V-D138 sequence of the CD9 antigen is the site which interacts with the HBD and may play an essential role in the upregulation of the juxtacrine activity of proHB-EGF.
肝素/硫酸乙酰肝素蛋白聚糖（HSPG）通过其由21个氨基酸残基组成的肝素结合域（HBD，即P21）与肝素结合表皮生长因子样生长因子（HB-EGF）结合。CD9抗原也与膜锚定形式的HB-EGF（前体HB-EGF）相互作用，并增强其旁分泌活性。CD9抗原增强前体HB-EGF和前体双调蛋白的旁分泌活性，但对前体转化生长因子-α没有影响。虽然HB-EGF和双调蛋白都含有一个HBD，但转化生长因子-α没有。这表明HB-EGF的HBD也参与CD9抗原的结合。缺乏HSPG的突变型中国仓鼠卵巢细胞在转染CD9抗原cDNA后恢复了与固定化P21结合的能力。这种结合被肝素以剂量依赖性方式竞争性抑制。利用表面等离子体共振分析了与CD9抗原细胞外结构域对应的合成肽与固定化P21之间的相互作用。CD9抗原的119VIKEVQEFYKDTYNKLKTKD138序列被认为代表HB-EGF的结合位点。肝素/P21和119V-D138/P21的解离常数（k(D)）值分别为(2.82±0.10)×10(-8)M和(3.71±0.71)×10(-5)M。这些结果表明，CD9抗原的119V-D138序列是与HBD相互作用的位点，可能在增强前体HB-EGF的旁分泌活性中起重要作用。
