title: HypE-specific nanobodies as tools to modulate HypE-mediated target AMPylation.
abstract: The covalent addition of mono-AMP to target proteins (AMPylation) by Fic domain-containing proteins is a poorly understood, yet highly conserved post-translational modification. Here, we describe the generation, evaluation, and application of four HypE-specific nanobodies: three that inhibit HypE-mediated target AMPylation in vitro and one that acts as an activator. All heavy chain-only antibody variable domains bind HypE when expressed as GFP fusions in intact cells. We observed localization of HypE at the nuclear envelope and further identified histones H2-H4, but not H1, as novel in vitro targets of the human Fic protein. Its role in histone modification provides a possible link between AMPylation and regulation of gene expression.
authors: From the Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142 and.
authors: Matthias C
authors: MC
authors: Truttmann
authors: From the Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142 and.
authors: Qin
authors: Q
authors: Wu
authors: From the Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142 and.
authors: Sarah
authors: S
authors: Stiegeler
authors: From the Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142 and.
authors: Joao N
authors: JN
authors: Duarte
authors: From the Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142 and.
authors: Jessica
authors: J
authors: Ingram
authors: From the Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142 and the Massachusetts Institute of Technology, Cambridge, Massachusetts 02139 ploegh@wi.mit.edu.
authors: Hidde L
authors: HL
authors: Ploegh
mesh_terms: D000249:Adenosine Monophosphate / Q000378:metabolism*; D000595:Amino Acid Sequence; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D002460:Cell Line; D005434:Flow Cytometry; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D009713:Nucleotidyltransferases; D017386:Sequence Homology, Amino Acid; D061905:Single-Domain Antibodies / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; D008565:Membrane Proteins; D061905:Single-Domain Antibodies; D000249:Adenosine Monophosphate; C541422:FICD protein, human; D009713:Nucleotidyltransferases
keywords: AMPylation; Fic Proteins; Histone Modification; HypE; Nuclear Membrane; Phage Display; Post-translational Modification (PTM); Recombinant Protein Expression; VHHs