scholarly journals Visualization of ligand-induced transmembrane signaling in the full-length human insulin receptor

2018 ◽  
Vol 217 (5) ◽  
pp. 1643-1649 ◽  
Author(s):  
Theresia Gutmann ◽  
Kelly H. Kim ◽  
Michal Grzybek ◽  
Thomas Walz ◽  
Ünal Coskun
Keyword(s):  
Insulin Receptor ◽  
Tyrosine Kinases ◽  
Structural Changes ◽  
Critical Role ◽  
Insulin Binding ◽  
Structural Rearrangement ◽  
Full Length ◽  
Transmembrane Signaling ◽  
Human Insulin Receptor ◽  
Multicellular Organisms

Insulin receptor (IR) signaling plays a critical role in the regulation of metabolism and growth in multicellular organisms. IRs are unique among receptor tyrosine kinases in that they exist exclusively as covalent (αβ)2 homodimers at the cell surface. Transmembrane signaling by the IR can therefore not be based on ligand-induced dimerization as such but must involve structural changes within the existing receptor dimer. In this study, using glycosylated full-length human IR reconstituted into lipid nanodiscs, we show by single-particle electron microscopy that insulin binding to the dimeric receptor converts its ectodomain from an inverted U-shaped conformation to a T-shaped conformation. This structural rearrangement of the ectodomain propagates to the transmembrane domains, which are well separated in the inactive conformation but come close together upon insulin binding, facilitating autophosphorylation of the cytoplasmic kinase domains.

scholarly journals Visualization of ligand-induced transmembrane signalling in the full-length human insulin receptor

2017 ◽  
Author(s):  
Theresia Gutmann ◽  
Kelly H. Kim ◽  
Michal Grzybek ◽  
Thomas Walz ◽  
Ünal Coskun
Keyword(s):  
Insulin Receptor ◽  
Human Insulin ◽  
Insulin Binding ◽  
Structural Rearrangement ◽  
Transmembrane Domains ◽  
Full Length ◽  
Transmembrane Signalling ◽  
Inactive Conformation ◽  
Human Insulin Receptor ◽  
Lipid Nanodiscs

ABSTRACTUsing glycosylated full-length human insulin receptor reconstituted into lipid nanodiscs, we show that insulin binding to the dimeric receptor converts its ectodomains from an inverted U-shaped to a T-shaped conformation. This unprecedented structural rearrangement of the ectodomains propagates to the transmembrane domains, which are well separated in the inactive conformation, but come together upon insulin binding, allowing autophosphorylation of the cytoplasmic kinase domains.

A Radioimmunoassay for Human Insulin Receptor Correlation Between Insulin Binding and Receptor Mass

1991 ◽  
Vol 23 (03) ◽  
pp. 117-121 ◽  
Author(s):  
G. Boden ◽  
F. Jadali ◽  
L. Tappy ◽  
Y. Fujita-Yamaguchi
Keyword(s):  
Insulin Receptor ◽  
Human Insulin ◽  
Insulin Binding ◽  
Human Insulin Receptor

scholarly journals Activation mechanism of the insulin receptor revealed by cryo-EM structure of the fully liganded receptor–ligand complex

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Emiko Uchikawa ◽  
Eunhee Choi ◽  
Guijun Shang ◽  
Hongtao Yu ◽  
Xiao-chen Bai
Keyword(s):  
Insulin Receptor ◽  
Insulin Signaling ◽  
Insulin Binding ◽  
Full Length ◽  
The Other ◽  
Activation Mechanism ◽  
Metabolic Homeostasis ◽  
Ligand Complex ◽  
Receptor Ligand ◽  
Cellular Assays

Insulin signaling controls metabolic homeostasis. Here, we report the cryo-EM structure of full-length insulin receptor (IR) and insulin complex in the active state. This structure unexpectedly reveals that maximally four insulins can bind the ‘T’-shaped IR dimer at four distinct sites related by 2-fold symmetry. Insulins 1 and 1’ bind to sites 1 and 1’, formed by L1 of one IR protomer and α-CT and FnIII-1 of the other. Insulins 2 and 2’ bind to sites 2 and 2’ on FnIII-1 of each protomer. Mutagenesis and cellular assays show that both sites 1 and 2 are required for optimal insulin binding and IR activation. We further identify a homotypic FnIII-2–FnIII-2 interaction in mediating the dimerization of membrane proximal domains in the active IR dimer. Our results indicate that binding of multiple insulins at two distinct types of sites disrupts the autoinhibited apo-IR dimer and stabilizes the active dimer.

Insulin Receptor: 3D Reconstruction from Darkfield Stem Images, Structural Interpretation and Functional Model

1999 ◽  
Vol 5 (S2) ◽  
pp. 408-409
Author(s):  
F.P. Ottensmeyer ◽  
R.Z.-T. Luo ◽  
A.B. Fernandes ◽  
D. Benia ◽  
C.C. Yip
Keyword(s):  
Growth Factor ◽  
Insulin Receptor ◽  
Tyrosine Kinases ◽  
Quaternary Structure ◽  
Functional Model ◽  
Growth Factor Receptor ◽  
Insulin Binding ◽  
Dark Field ◽  
Bovine Insulin ◽  
Structural Basis

We have reconstructed the three-dimensional quaternary structure of the complete 480 kDa insulin receptor (IR), complexed with NanoGold-labelled insulin, via sets of electron micrographs obtained by low-dose low-temperature dark field scanning transmission electron microscopy (STEM).Insulin binding to IR in mammalian cell membranes is essential for its manifold effects such as glucose homeostasis, increased protein synthesis, growth, and development. IR belongs to the superfamily of transmembrane receptor tyrosine kinases that include the monomeric epidermal growth factor receptor (EGFR) and platelet-derived growth factor receptor (PDGFR). In contrast, IR and its homologues IGF-1R (insulin-like growth factor 1 receptor) and IRR (insulin receptorrelated receptor) are sub-types of this family that are intrinsic disulfide-linked dimers of two αβ heterodimers. Monomeric receptor TKs are inactive, but are activated by ligand-induced dimerization that results in autophosphorylation. IR-like TKs are also inactive even though they are already dimeric, and are activated by ligand binding without further oligomerization. Insulin binding to the extracellular domain of IR results in autophosphorylation of specific tyrosines to initiate an intracellular signal transduction cascade. However, because the quaternary structure of IR is not known, the structural basis for the mechanism of IR activation by extracellular insulin binding has not been elucidated.The insulin receptor was purified from human placenta. Bovine insulin was derivatized with NanoGold at the B chain Phel, a location not directly involved in receptor binding. Binding of derivatized insulin to the purified receptor was reduced only slightly compared to binding of the native insulin.

scholarly journals Secretion of the extracellular domain of the human insulin receptor from insect cells by use of a baculovirus vector

1989 ◽  
Vol 261 (1) ◽  
pp. 119-126 ◽  
Author(s):  
J Sissom ◽  
L Ellis
Keyword(s):  
Insulin Receptor ◽  
Human Insulin ◽  
Mammalian Cells ◽  
Culture Medium ◽  
Insect Cells ◽  
Transmembrane Domain ◽  
Binding Protein ◽  
Insulin Binding ◽  
Cell System ◽  
Human Insulin Receptor

To explore the utility of the baculovirus/insect-cell system for the expression of a soluble secreted human insulin-receptor (hIR) extracellular ligand-binding domain, we have engineered a recombinant virus encoding an hIR deletion mutant which is truncated eight residues from the beginning of the predicted transmembrane domain (i.e. 921 residues). Within 24 h after infection of Sf9 cells with virus, insulin-binding activity begins to accumulate in the culture medium, and reaches a maximum between 48 and 72 h. The intracellular transit and processing of this secreted receptor, designated ‘AchIR01’, is quite slow. After 24 h in pulse-chase experiments approximately 50% of the metabolically labelled protein is still inside the cell. This protein accumulates as a non-cleaved hIR precursor which is glycosylated, but the carbohydrate is entirely endoglycosidase H (endoH)-sensitive (i.e. high mannose). Approximately one-half of the receptor in the culture medium (i.e. approximately 25% of the total) is in the form of non-cleaved precursor, and about one half of its carbohydrate chains are now endoH-resistant. The remainder of the protein is proteolytically processed hIR (alpha-plus truncated beta-subunits). None of these hIR species exhibit O-linked carbohydrate. Only the processed form of the receptor in the medium binds insulin. This insulin-binding protein is secreted as a dimer (alpha beta)2, and binds insulin with an affinity which is comparable with that of both the wild-type hIR as well as the secreted form of the hIR expressed in mammalian cells. Despite the rather inefficient processing and altered glycosylation of the AchIR01 protein in insect cells, this high-affinity insulin-binding protein accumulates in the medium at levels (mg/litre) of about 100 times that achieved in a mammalian-cell system.

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