Biochemical Studies of the ER Resident Kinase PERK and Implications for Local Translational Control
type: CreativeWork
description: The unfolded protein response (UPR) is a highly conserved protein quality control mechanism that becomes activated in response to misfolded protein accumulation in the endoplasmic reticulum (ER) resulting in ER stress. The ER resident kinase protein kinase R (PKR)-like endoplasmic reticulum kinase (PERK) is able to couple ER stress to translational repression and transcriptional reprogramming through phosphorylation of eIF2α. This is achieved through the recruitment of eIF2α by a unique cytoplasmic loop in PERK’s kinase domain. PERK signalling has been shown to play an important role in disease pathology notably in neurodegenerative diseases. Increased levels of phosphorylated PERK and eIF2α have been observed in the brains of patients with Alzheimer’s disease, Parkinson’s disease, other tauopathies, and prion disease. In many of these conditions, the associated disease-specific misfolded proteins accumulate in the cytoplasm instead of the ER. Currently, the mechanism for PERK activation by cytoplasmic misfolded proteins, as opposed to those in the ER, remains elusive. To gain insight into novel aspects of cytoplasmic PERK signalling which may be relevant to cellular health and disease, in particular, neurodegenerative disease, the PERK-protein interactome was examined using mass spectrometry. This led to the identification of the mitochondrial protein, ATAD3A, as a PERK kinase insert loop interactor. Using biochemical and biophysical techniques, a common binding site within the kinase insert loop of PERK was identified for eIF2α and ATAD3A. To further investigate the dynamics of these interactions bioluminescence resonance energy transfer (BRET) based biosensors were developed, enabling the interaction between PERK and eIF2α to be monitored in real-time in live cells. Mutating the kinase insert loop supported the in vitro binding assays by also indicating the presence of two eIF2α binding sites within PERK’s loop. Furthermore, ATAD3A can outcompete eIF2α for occupa…
abstract: The unfolded protein response (UPR) is a highly conserved protein quality control mechanism that becomes activated in response to misfolded protein accumulation in the endoplasmic reticulum (ER) resulting in ER stress. The ER resident kinase protein kinase R (PKR)-like endoplasmic reticulum kinase (PERK) is able to couple ER stress to translational repression and transcriptional reprogramming through phosphorylation of eIF2α. This is achieved through the recruitment of eIF2α by a unique cytoplasmic loop in PERK’s kinase domain. PERK signalling has been shown to play an important role in disease pathology notably in neurodegenerative diseases. Increased levels of phosphorylated PERK and eIF2α have been observed in the brains of patients with Alzheimer’s disease, Parkinson’s disease, other tauopathies, and prion disease. In many of these conditions, the associated disease-specific misfolded proteins accumulate in the cytoplasm instead of the ER. Currently, the mechanism for PERK activation by cytoplasmic misfolded proteins, as opposed to those in the ER, remains elusive. To gain insight into novel aspects of cytoplasmic PERK signalling which may be relevant to cellular health and disease, in particular, neurodegenerative disease, the PERK-protein interactome was examined using mass spectrometry. This led to the identification of the mitochondrial protein, ATAD3A, as a PERK kinase insert loop interactor. Using biochemical and biophysical techniques, a common binding site within the kinase insert loop of PERK was identified for eIF2α and ATAD3A. To further investigate the dynamics of these interactions bioluminescence resonance energy transfer (BRET) based biosensors were developed, enabling the interaction between PERK and eIF2α to be monitored in real-time in live cells. Mutating the kinase insert loop supported the in vitro binding assays by also indicating the presence of two eIF2α binding sites within PERK’s loop. Furthermore, ATAD3A can outcompete eIF2α for occupa…
author: Brar, Karinder
language: eng
keywords: biology
license: https://www.repository.cam.ac.uk/bitstreams/31eb194d-6d1d-4196-9b56-15094c3f29bb/download
license: https://www.rioxx.net/licenses/all-rights-reserved/
date published: 2023-09-29
date modified: 2026-07-22T22:23:57Z
provider: Cambridge University
publisher: Networked Digital Library of Theses and Dissertations
type: WebPage
source organization: University of Cambridge

PropertyValue
property ID: DOI
value: 10.17863/CAM.108433