The intramolecular domain interactions and phosphatase activation mechanisms of Shp2
type: CreativeWork
description: Shp2 (Src Homology 2 Domain-containing Protein Tyrosine Phosphatase 2) is a pivotal player in various signaling pathways in response to growth factors. It contains two SH2 domains (N-SH2 and C-SH2), a phosphatase catalytic domain and a C-terminal tail containing two tyrosyl phosphorylation sites. While the tyrosine phosphorylation of C-terminal tail has been shown to be critical for the proper Shp2 functions, there is a lack of understanding on how the phosphorylation of these two tyrosine sites regulates the conformational changes and functions of Shp2. Here, we developed a FRET-based Shp2 reporter, a single molecule comprised of the full length human Shp2 concatenated in between a FRET pair, ECFP and YPet. In vitro incubation of Shp2 reporter with PDGFRβ induced a significant FRET increase via intramolecular interaction between C-SH2 and phosphotyrosine 580 (pY580). The phosphotyrosine 542 (pY542) has an adverse position despite a favored surrounding amino acid sequence such that it can only bind to C-SH2 in the absence of pY580. In contrast, pY542 dominates the binding of C-SH2 in mouse embryonic fibroblasts (MEFs). This intramolecular binding is essential for the subsequent effective phosphorylation of Y580, which is then sequestered by Grb2 in cells. These results suggested that the cellular environment can tune and switch the interaction of C-SH2 toward pY542. When the sequences containing Y542 and Y580 were engineered to exchange their positions such that the favored sequence containing Y542 is located at the advantageous position of 580 (Y542’), pY542’ constitutively dominates the binding of C-SH2 both in vitro and in MEFs. This rewiring of intramolecular interactions within Shp2 also caused a reprogramming of downstream signaling events, resulting in a more transient ERK phosphorylation. Therefore, our results indicate that the combination of a favorable sequence with an adverse position can create a delicate balance and plasticity of intramolecular interac…
abstract: Shp2 (Src Homology 2 Domain-containing Protein Tyrosine Phosphatase 2) is a pivotal player in various signaling pathways in response to growth factors. It contains two SH2 domains (N-SH2 and C-SH2), a phosphatase catalytic domain and a C-terminal tail containing two tyrosyl phosphorylation sites. While the tyrosine phosphorylation of C-terminal tail has been shown to be critical for the proper Shp2 functions, there is a lack of understanding on how the phosphorylation of these two tyrosine sites regulates the conformational changes and functions of Shp2. Here, we developed a FRET-based Shp2 reporter, a single molecule comprised of the full length human Shp2 concatenated in between a FRET pair, ECFP and YPet. In vitro incubation of Shp2 reporter with PDGFRβ induced a significant FRET increase via intramolecular interaction between C-SH2 and phosphotyrosine 580 (pY580). The phosphotyrosine 542 (pY542) has an adverse position despite a favored surrounding amino acid sequence such that it can only bind to C-SH2 in the absence of pY580. In contrast, pY542 dominates the binding of C-SH2 in mouse embryonic fibroblasts (MEFs). This intramolecular binding is essential for the subsequent effective phosphorylation of Y580, which is then sequestered by Grb2 in cells. These results suggested that the cellular environment can tune and switch the interaction of C-SH2 toward pY542. When the sequences containing Y542 and Y580 were engineered to exchange their positions such that the favored sequence containing Y542 is located at the advantageous position of 580 (Y542’), pY542’ constitutively dominates the binding of C-SH2 both in vitro and in MEFs. This rewiring of intramolecular interactions within Shp2 also caused a reprogramming of downstream signaling events, resulting in a more transient ERK phosphorylation. Therefore, our results indicate that the combination of a favorable sequence with an adverse position can create a delicate balance and plasticity of intramolecular interac…
author: Sun, Jie
language: en
keywords: Shp2 (Src Homology 2 Domain-containing Protein Tyrosine Phosphatase 2), FRET reporter, Intramolecular interactions, Phosphatase activation, Fluorescent/ Förster resonance energy transfer (FRET)
date published: 2012-02-01T00:53:22Z
date modified: 2026-07-22T22:25:27Z
provider: University of Illinois - Urbana-Champaign
publisher: Networked Digital Library of Theses and Dissertations
type: WebPage
source organization: University of Illinois at Urbana-Champaign

PropertyValue
property ID: Handle
value: http://hdl.handle.net/2142/29512