UT-BIORAD-FARM CR Casaccia ENEA, Rome, Italy.
PLoS One. 2012;7(2):e32555. doi: 10.1371/journal.pone.0032555. Epub 2012 Feb 28.
Protein tyrosine phosphatase ρ (PTPρ) belongs to the classical receptor type IIB family of protein tyrosine phosphatase, the most frequently mutated tyrosine phosphatase in human cancer. There are evidences to suggest that PTPρ may act as a tumor suppressor gene and dysregulation of Tyr phosphorylation can be observed in diverse diseases, such as diabetes, immune deficiencies and cancer. PTPρ variants in the catalytic domain have been identified in cancer tissues. These natural variants are nonsynonymous single nucleotide polymorphisms, variations of a single nucleotide occurring in the coding region and leading to amino acid substitutions. In this study we investigated the effect of amino acid substitution on the structural stability and on the activity of the membrane-proximal catalytic domain of PTPρ. We expressed and purified as soluble recombinant proteins some of the mutants of the membrane-proximal catalytic domain of PTPρ identified in colorectal cancer and in the single nucleotide polymorphisms database. The mutants show a decreased thermal and thermodynamic stability and decreased activation energy relative to phosphatase activity, when compared to wild- type. All the variants show three-state equilibrium unfolding transitions similar to that of the wild- type, with the accumulation of a folding intermediate populated at ~4.0 M urea.
蛋白酪氨酸磷酸酶 ρ（PTPρ）属于经典的受体型 IIB 家族的蛋白酪氨酸磷酸酶，是人类癌症中最常突变的酪氨酸磷酸酶。有证据表明 PTPρ 可能作为肿瘤抑制基因发挥作用，并且在多种疾病中，如糖尿病、免疫缺陷和癌症中，可以观察到 Tyr 磷酸化的失调。在癌症组织中已经鉴定出催化结构域中的 PTPρ 变体。这些天然变体是非同义单核苷酸多态性，即编码区域中单个核苷酸的变异，导致氨基酸取代。在这项研究中，我们研究了氨基酸取代对 PTPρ 膜近端催化结构域的结构稳定性和活性的影响。我们表达和纯化了一些在结直肠癌和单核苷酸多态性数据库中鉴定出的 PTPρ 膜近端催化结构域的突变体作为可溶性重组蛋白。与野生型相比，突变体显示出热稳定性和热力学稳定性降低，并且相对于磷酸酶活性的活化能降低。所有变体均显示出与野生型相似的三态平衡展开转变，在~4.0 M 脲中积累了一个折叠中间体。
