Abstract
Placental malfunction induces pregnancy disorders which contribute to life-threatening complications for both the mother and the fetus. Identification and characterization of placental multi-protein complexes is an important step to integratedly understand the protein-protein interaction networks in placenta which determine placental function. In this study, blue native/sodium dodecyl sulfate polyacrylamide gel electrophoresis (BN/SDS-PAGE) and Liquid chromatography-tandem mass spectrometry (LC-MS/MS) were used to screen the multi-protein complexes in placenta. 733 unique proteins and 34 known and novel heterooligomeric multi-protein complexes including mitochondrial respiratory chain complexes, integrin complexes, proteasome complexes, histone complex, and heat shock protein complexes were identified. A novel protein complex, which involves clathrin and small conductance calcium-activated potassium (SK) channel protein 2, was identified and validated by antibody based gel shift assay, co-immunoprecipitation and immunofluorescence staining. These results suggest that BN/SDS-PAGE, when integrated with LC-MS/MS, is a very powerful and versatile tool for the investigation of placental protein complexes. This work paves the way for deeper functional characterization of the placental protein complexes associated with pregnancy disorders.
Highlights
The placenta plays a pivotal role of promoting the exchange of nutrients and waste products between the maternal and fetal circulatory systems [1]
Those studies have identified a number of abnormally expressed proteins in plasma, amniotic fluid, intact placentae, or trophoblasts from preeclampsia patients using various proteomic techniques including classical two dimensional (2D) gel electrophoresis, Isobaric tags for relative and absolute quantitation, and Difference gel electrophoresis (DIGE) [5]
We report using Blue native PAGE (BN-PAGE) to identify and characterize a number of functional protein complexes from placenta, which supplements a new methodology for the proteomic analysis of placenta
Summary
The placenta plays a pivotal role of promoting the exchange of nutrients and waste products between the maternal and fetal circulatory systems [1]. Maternal preeclampsia (PE) and fetal intrauterine growth restriction (IUGR) are two of the most common and serious complications of human pregnancy associated with placental abnormalities [2,3]. Development and functionalization of placenta are mediated by various proteins which have been investigated in proteomics and disciplines associated. Those studies have identified a number of abnormally expressed proteins in plasma, amniotic fluid, intact placentae, or trophoblasts from preeclampsia patients using various proteomic techniques including classical two dimensional (2D) gel electrophoresis, Isobaric tags for relative and absolute quantitation (iTRAQ), and Difference gel electrophoresis (DIGE) [5]. They are not able to provide information about how these proteins interact with each other
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