Dr. Reddy's Institute of Life Sciences, University of Hyderabad, Hyderabad 500046, India.
Proc Natl Acad Sci U S A. 2013 Dec 17;110(51):20467-72. doi: 10.1073/pnas.1306145110. Epub 2013 Nov 26.
Resistin, a cysteine-rich adipocytokine, proposed as a link between obesity and diabetes in mice, was shown as a proinflammatory molecule in humans. We earlier reported that human resistin (hRes), a trimer, was resistant to heat and urea denaturation, existed in an oligomeric polydispersed state, and showed a concentration-dependent conformational change. These properties and an intimate correlation of hRes expression with cellular stress prompted us to investigate hRes as a possible chaperone. Here, we show that recombinant human resistin was able to protect the heat-labile enzymes citrate synthase and Nde1 from thermal aggregation and inactivation and was able to refold and restore their enzymatic activities after heat/guanidinium chloride denaturation. Furthermore, recombinant human resistin could bind misfolded proteins only. Molecular dynamics-based association-dissociation kinetics of hRes subunits pointed to resistin being a molecular chaperone. Bis-ANS, which blocks surface hydrophobicity, abrogated the chaperone activity of hRes, establishing the importance of surface hydrophobicity for chaperone activity. Replacement of Phe49 with Tyr (F49YhRes), a critical residue within the hydrophobic patch of hRes, although it could prevent thermal aggregation of citrate synthase and Nde1, was unable to refold and restore their activities. Treatment of U937 cells with tunicamycin/thapsigargin resulted in reduced hRes secretion and concomitant localization in the endoplasmic reticulum. Escherichia coli transformants expressing hRes could be rescued from thermal stress, pointing to hRes's chaperone-like function in vivo. HeLa cells transfected with hRes showed protection from thapsigargin-induced apoptosis. In conclusion, hRes, an inflammatory protein, additionally exhibited chaperone-like properties, suggesting a possible link between inflammation and cellular stress.
抵抗素是一种富含半胱氨酸的脂肪细胞因子，在小鼠中被认为是肥胖与糖尿病之间的联系，在人类中被证明是一种促炎分子。我们之前报道过，人类抵抗素（hRes）是一种三聚体，对热和尿素变性具有抗性，存在于多分散的寡聚状态，并表现出浓度依赖性的构象变化。这些特性以及 hRes 表达与细胞应激之间的密切关系促使我们将 hRes 作为一种潜在的伴侣蛋白进行研究。在这里，我们证明重组人抵抗素能够保护不耐热的酶柠檬酸合酶和 Nde1 免受热聚集和失活，并能够在热/盐酸胍变性后重新折叠并恢复其酶活性。此外，重组人抵抗素只能结合错误折叠的蛋白质。基于分子动力学的 hRes 亚基的结合-解离动力学表明抵抗素是一种分子伴侣。结合阻止表面疏水性的双-ANS，消除了 hRes 的伴侣活性，从而证明了表面疏水性对于伴侣活性的重要性。用 Tyr（F49YhRes）取代 Phe49（hRes 疏水斑内的关键残基），尽管可以防止柠檬酸合酶和 Nde1 的热聚集，但不能重新折叠并恢复其活性。用衣霉素/他普西龙处理 U937 细胞会导致 hRes 分泌减少，并伴有内质网定位。表达 hRes 的大肠杆菌转化体可以从热应激中得到挽救，这表明 hRes 在体内具有伴侣样功能。转染 hRes 的 HeLa 细胞显示出对他普西龙诱导的细胞凋亡的保护。总之，作为一种炎症蛋白的 hRes 还表现出伴侣样特性，这表明炎症和细胞应激之间可能存在联系。
