The C-terminal phosphorylation domain of the epidermal growth factor receptor is believed to regulate protein kinase activity as well as mediate the assembly of signal transduction complexes. The structure and dynamics of this proposed autoregulatory domain were examined by labeling the extreme C terminus of the EGFR intracellular domain (ICD) with an extrinsic fluorophore. Fluorescence anisotropy decay analysis of the nonphosphorylated EGFR-ICD yielded two rotational correlation times: a longer time, consistent with the global rotational motion of a 60- to 70-kDa protein with an elongated globular conformation, and a shorter time, presumably contributed by segmental motion near the fluorophore. A C-terminally truncated form of EGFR-ICD yielded a slow component consistent with the rotational motion of the 38-kDa kinase core. These findings suggested a structural arrangement of the EGFR-ICD in which the C-terminal phosphorylation domain interacts with the kinase core to move as an extended structure. A marked reduction in the larger correlation time of EGFR-ICD was observed upon its autophosphorylation. This dynamic component was faster than predicted for the globular motion of the 62-kDa EGFR-ICD, suggesting an increase in the mobility of the C-terminal domain and a likely displacement of this domain from the kinase core. The interaction between the SH2 domain of c-Src and the phosphorylated EGFR C-terminal domain was shown to impede its mobility. Circular dichroism spectroscopy indicated that the EGFR C-terminal domain possessed a significant level of secondary structure in the form of alpha-helices and beta-sheets, with a marginal change in beta-sheet content occurring upon phosphorylation.
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http://dx.doi.org/10.1110/ps.052045306 | DOI Listing |
Nat Struct Mol Biol
January 2025
Department of Biochemistry and Molecular Cell Biology, Key Laboratory of Cell Differentiation and Apoptosis of Chinese Ministry of Education, Shanghai Key Laboratory for Tumor Microenvironment and Inflammation, Shanghai Jiao Tong University School of Medicine, Shanghai, China.
The Rpd3S histone deacetylase complex has a crucial role in genomic integrity by deacetylating transcribed nucleosomes following RNA polymerase (Pol) II passage. Cryo-EM studies highlight the importance of asymmetrical Rco1-Eaf3 dimers in nucleosome binding, yet the interaction dynamics with nucleosomal substrates alongside elongating Pol II are poorly understood. Here we demonstrate the essential function of the Rco1 N-terminal intrinsically disordered region (IDR) in modulating Pol II association, in which K/R mutations within the Rco1 IDR impair interaction of Rpd3S with the C-terminal domain (CTD) of Rpb1, without affecting nucleosome recognition or complex integrity.
View Article and Find Full Text PDFNat Commun
January 2025
Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA.
Members of the piggyBac superfamily of DNA transposons are widely distributed in host genomes ranging from insects to mammals. The human genome has retained five piggyBac-derived genes as domesticated elements although they are no longer mobile. Here, we have investigated the transposition properties of piggyBat from Myotis lucifugus, the only known active mammalian DNA transposon, and show that its low activity in human cells is due to subterminal inhibitory DNA sequences.
View Article and Find Full Text PDFBiochem Biophys Rep
March 2025
College of Biomedical Sciences, Larkin University, Miami, FL, 33169, USA.
J Biol Chem
December 2024
Laboratory of Molecular Microbiology and Genetics, BRIC-Centre for DNA Fingerprinting and Diagnostics, Hyderabad, India. Electronic address:
Genetic studies in Escherichia coli have implicated the unphosphorylated version of PtsN (unphospho-PtsN), the terminal phospho-acceptor of the PtsP-PtsO-PtsN phosphorelay, as a negative regulator of potassium (K) efflux mediated by YcgO. YcgO is a protein belonging to the CPA1 family of monovalent cation/proton antiporters. Here we show that in vivo, YcgO comprises an approximately 383 amino acid N-terminal transmembrane domain (TMD) and a 195 amino acid C-terminal cytoplasmic region (CTR).
View Article and Find Full Text PDFNucleic Acids Res
December 2024
Department of Biology, Massachusetts Institute of Technology, Building 68, 31 Ames St., Cambridge, MA 02139, USA.
The eukaryotic microrchidia (MORC) protein family are DNA gyrase, Hsp90, histidine kinase, MutL (GHKL)-type ATPases involved in gene expression regulation and chromatin compaction. The molecular mechanisms underlying these activities are incompletely understood. Here, we studied the full-length human MORC2 protein biochemically.
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