Department of Biochemistry, Medical College of Wisconsin, Milwaukee, WI, USA.
Proteomics. 2011 Apr;11(8):1382-90. doi: 10.1002/pmic.201000362. Epub 2011 Mar 1.
Maspin, a 42-kDa non-classical serine protease inhibitor (serpin), is expressed by epithelial cells of various tissues including the cornea. The protein localizes to the nucleus and cytosol, and is present in the extracellular space. While extracellular maspin regulates corneal stromal fibroblast adhesion and inhibits angiogenesis during wound healing in the cornea, the molecular mechanism of its extracellular functions is unclear. We hypothesized that identifying post-translational modifications of maspin, such as phosphorylation, may help decipher its mode of action. The focus of this study was on the identification of phosphorylation sites on extracellular maspin, since the extracellular form of the molecule is implicated in several functions. Multi-stage fragmentation MS was used to identify sites of phosphorylation on extracellular corneal epithelial cell maspin. A total of eight serine and threonine phosphorylation sites (Thr50, Ser97, Thr118, Thr157, Ser240, Ser298, Thr310 and Ser316) were identified on the extracellular forms of the molecule. Phosphorylation of tyrosine residues was not detected on extracellular maspin from corneal epithelial cell, in contrast to breast epithelial cells. This study provides the basis for further investigation into the functional role of phosphorylation of corneal epithelial maspin.
Maspin 是一种 42kDa 的非经典丝氨酸蛋白酶抑制剂（serpin），存在于多种组织的上皮细胞中，包括角膜。该蛋白定位于细胞核和细胞质，也存在于细胞外基质中。虽然细胞外的 maspin 可以调节角膜基质成纤维细胞的黏附，并在角膜伤口愈合过程中抑制血管生成，但它的细胞外功能的分子机制尚不清楚。我们假设鉴定 maspin 的翻译后修饰（如磷酸化）可能有助于阐明其作用机制。本研究的重点是鉴定细胞外 maspin 的磷酸化位点，因为该分子的细胞外形式与多种功能有关。采用多级碎裂 MS 技术鉴定了角膜上皮细胞外 maspin 的磷酸化位点。在该分子的细胞外形式上共鉴定到 8 个丝氨酸和苏氨酸磷酸化位点（Thr50、Ser97、Thr118、Thr157、Ser240、Ser298、Thr310 和 Ser316）。与乳腺上皮细胞不同，在角膜上皮细胞的细胞外 maspin 中未检测到酪氨酸残基的磷酸化。本研究为进一步研究角膜上皮细胞 maspin 磷酸化的功能作用提供了基础。
