Structure of a Numb PTB domain–peptide complex suggests a basis for diverse binding specificity

Li, SC; Zwahlen, Catherine; Sj, Vincent; Cj, McGlade; Le, Kay; Pawson, Tony; Forman-Kay, Julie D.

doi:10.1038/4185

Cited by 111 publications

(113 citation statements)

References 48 publications

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“…Although most PTB domains that have been solved share low levels of sequence identity, structure-based alignments reveal that the core topological structure is evolutionary conserved (19). Like the PTB domains of Shc, disabled, and numb, the Mint1 PTB domain contains an N-terminal helix (α1) inserted between strands β1 and β2 (17,(20)(21)(22). Interestingly, our structure shows that the linker region C-terminal to the PTB domain forms a short helix (α3) that folds back onto the core structure of the PTB* domain and sterically hinders the APP binding site (Fig.…”

Section: Resultsmentioning

confidence: 99%

Autoinhibition of Mint1 adaptor protein regulates amyloid precursor protein binding and processing

Matos

Xu²,

Dulubova

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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Mint adaptor proteins bind to the amyloid precursor protein (APP) and regulate APP processing associated with Alzheimer's disease; however, the molecular mechanisms underlying Mint regulation in APP binding and processing remain unclear. Biochemical, biophysical, and cellular experiments now show that the Mint1 phosphotyrosine binding (PTB) domain that binds to APP is intramolecularly inhibited by the adjacent C-terminal linker region. The crystal structure of a C-terminally extended Mint1 PTB fragment reveals that the linker region forms a short α-helix that folds back onto the PTB domain and sterically hinders APP binding. This intramolecular interaction is disrupted by mutation of Tyr633 within the Mint1 autoinhibitory helix leading to enhanced APP binding and β-amyloid production. Our findings suggest that an autoinhibitory mechanism in Mint1 is important for regulating APP processing and may provide novel therapies for Alzheimer's disease.M ints [also known as X11s or amyloid precursor protein (APP) binding, family A] are a family of adaptor proteins encoded by three genes: neuron-specific Mint1 and Mint2, and the ubiquitously expressed Mint3 (1, 2). Mints consist of a divergent N-terminal region and conserved C-terminal sequences composed of a phosphotyrosine binding (PTB) domain and two tandem PDZ (postsynaptic density-95/discs large/zona occludens-1) domains. The PTB domain of all Mints interacts directly with a conserved YENPTY cytoplasmic motif of the APP that is essential for regulating APP trafficking and processing (3-6). APP is a type I transmembrane protein that is sequentially cleaved by site-specific proteases (7-10). Abnormal processing of APP generates Aβ peptides leading to the accumulation of extracellular plaques that are characteristic of Alzheimer's disease (AD). β-Secretase is the first amyloidogenic cleavage, generating a soluble APP ectodomain (sAPPβ) and a C-terminal fragment (CTFβ). CTFβ is subsequently cleaved by γ-secretase (a protein complex of anterior pharyx defective 1, nicastrin, presenilin, and presenilin enhancer 2) to release the small peptides Aβ40 and Aβ42, the latter having an increased propensity to form amyloid aggregates found in the brains of AD patients (11)(12)(13)(14). Alternatively, APP is sequentially cleaved by α-secretase within the Aβ region then γ-secretase, which initiates the nonamyloidogenic or nontoxic pathway (9, 10).Considerable evidence indicates that the APP-Mint interaction is biologically significant because loss of each individual Mint protein delays age-dependent production of amyloid plaques in transgenic AD mouse models (15). Past studies have indicated that the N-and C-terminal regions of Mints play an important role in regulating APP binding and cleavage events based on deletion analysis of Mints in a heterologous expression system (6, 16). The Mint N terminus was shown to promote APP binding and Aβ production, whereas the C-terminal region disrupted PTB-mediated binding to APP and decreased Aβ peptides (6, 16). However, the structural basis ...

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Section: Resultsmentioning

confidence: 99%

Autoinhibition of Mint1 adaptor protein regulates amyloid precursor protein binding and processing

Matos

Xu²,

Dulubova

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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“…In the structure of IRS-1 bound to the IR peptide, the side chain of Tyr(P) 1009 of the IR buries the aliphatic portion of the structurally homologous Arg 212 side chain and hydrogen bonds to its guanido group (35). By contrast, the PTB domains of X11, Numb, and Shc contain a Ser at the homologous position (33,49), which is a more typical residue in PTB domains. In agreement with the prediction of its functional importance, mutation of Ile 139 of Icap1␣ to alanine abolished the Icap1␣-␤ 1 peptide interaction (Fig.…”

Section: A Mutational Analysis Of Icap1␣ Supports the Structural Predmentioning

confidence: 99%

Molecular Basis for Interaction between Icap1α PTB Domain and β1 Integrin

Chang¹,

Hoang²,

Liu³

et al. 2002

Journal of Biological Chemistry

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Icap1␣ is a 200-amino acid protein that binds to theIntegrins are transmembrane heterodimeric receptors for extracellular matrix and cell surface proteins (1, 2). The binding of integrins to ligands in the extracellular matrix is linked to cell attachment and spreading, which in turn activates various cytosolic signal cascades to promote cell migration, survival, proliferation, and differentiation (3-5). The binding of integrins to their ligands requires integrins to be in an "activated" conformation (6). The regulation of the activation status of integrins and the post-ligand binding activation of various signaling cascades require the integrin cytoplasmic domains. Mutagenesis studies have shown that the cytoplasmic domain of ␤ subunit is important for cell adhesion and migration (7,8) and for localization of integrins to focal contacts (9, 10).Integrin ␤ subunit cytoplasmic domain interacts with several cytoskeletal proteins, including ␣-actinin, talin, paxillin, and filamin (11-16), which may localize integrins to the site of cell-extracellular matrix interaction. Signaling molecules, such as FAK, ILK-1 (17), RACK1 (18), and Shc (19,20), also bind to the integrin ␤ subunit cytoplasmic domains and may link integrins directly to the cytosolic kinase cascades. In addition to these proteins, which can bind to the cytoplasmic domains of several different ␤ subunits, there are proteins with binding specificity toward a particular ␤ integrin cytoplasmic domain. For example, Cytohesin-1 interacts specifically with the ␤ 2 integrin (21) and regulates the leukocyte-specific ␣ L ␤ 2 integrins (22). Similarly, ␤ 3 -endonexin binds to the ␤ 3 integrin (23) and may play a role in the affinity regulation of platelet integrin ␣ IIB ␤ 3 (24). Another of these proteins is Icap1␣, 1 which displays a restricted binding toward the ␤ 1 integrins (25).Although the precise function of Icap1␣ has not been established, its role in integrin-dependent cell adhesion was suggested from the finding that Icap1␣ undergoes an adhesion-dependent phosphorylation (25). Furthermore, expression of a mutant Icap1␣ with a T38D mutation interferes with cell spreading (26). The Icap1␣ binding site on the ␤ 1 integrin has been mapped to the COOH-terminal region of ␤ 1 integrin (25), which includes one of the two NPXY (or NXXY) motifs present in several integrin ␤ subunits. These NPXY motifs are important for the localization of integrins to focal contacts (10), integrin-mediated endocytosis (27), and affinity regulation of integrins (28). The functions of ␤ 1 and ␤ 3 integrins can also be regulated by tyrosine phosphorylation at the NPXY motifs (29 -31).Here, we examined the structural basis for the interaction between Icap1␣ and the integrin ␤ 1 cytoplasmic domain. Sequence homology and molecular modeling reveal that Icap1␣ has a PTB fold. The PTB domains bind their target peptides through extensive hydrogen bonding and packing interactions in which the target peptide is bound to a characteristic grove on the PTB domain. Furthermore, some PTB domains ...

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“…Determination of a solution structure for a peptide⅐PTB domain complex and peptide binding studies indicate that the prototypic Drosophila Numb PTB module interacts with multiple target proteins by providing a unitary but flexible binding surface (50,51). This surface accommodates several classes of structurally diverse peptides that are presented in either type I ␤-turn or ␣-helical turn conformations (50,51). Analogies between Numb and CKA1 PTB domains strongly suggest that the cited binding properties will be evident in the PKC3-docking protein.…”

Section: Resultsmentioning

confidence: 99%

“…Numb proteins mediate asymmetric localization of several regulatory proteins in vivo (52,53). Determination of a solution structure for a peptide⅐PTB domain complex and peptide binding studies indicate that the prototypic Drosophila Numb PTB module interacts with multiple target proteins by providing a unitary but flexible binding surface (50,51). This surface accommodates several classes of structurally diverse peptides that are presented in either type I ␤-turn or ␣-helical turn conformations (50,51).…”

Section: Resultsmentioning

confidence: 99%

A Novel Adapter Protein Employs a Phosphotyrosine Binding Domain and Exceptionally Basic N-terminal Domains to Capture and Localize an Atypical Protein Kinase C

Zhang

Wu²,

Rubin

2001

Journal of Biological Chemistry

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Atypical protein kinase C isoforms (aPKCs) transmit regulatory signals to effector proteins located in the cytoplasm, nucleus, cytoskeleton, and membranes. Mechanisms by which aPKCs encounter and control effector proteins in various microenvironments are poorly understood. By using a protein interaction screen, we discovered two novel proteins that adapt a Caenorhabditis elegans aPKC (PKC3) for specialized (localized) functions; protein kinase C adapter 1 (CKA1, 593 amino acids) and CKA1S (549 amino acids) are derived from a unique mRNA by alternative utilization of two translation initiation codons. CKA1S and CKA1 are routed to the cell periphery by exceptionally basic N-terminal regions that include classical phosphorylation site domains (PSDs). Many hormones and growth factors elicit activation of phospholipases that produce diacylglycerol (DAG) 1 (1, 2). Protein kinase C (PKC) isoenzymes disseminate signals carried by DAG. Amplification and routing of signals are achieved because classical (␣, ␤I, ␤II, ␥) and novel (␦, ⑀, , , ) PKC isoforms (cPKCs and nPKCs, respectively) are DAG-dependent Ser/Thr phosphotransferases that translocate from cytoplasm to sites in membranes where DAG accumulates in response to hormones (1-4). Activated PKCs phosphorylate proteins that control secretion, mitogenesis, cytoskeleton organization, gene transcription, and many other physiological processes (1-3, 5-9).Atypical PKCs (aPKCs), which include vertebrate PKC and PKC isoforms and Caenorhabditis elegans PKC3 (1, 4, 8), also regulate critical cell functions. PKC and/or PKC activate the Ras-mitogen-activated protein kinase cascade, stimulate gene transcription, inhibit apoptosis, modulate ion channel activities, phosphorylate nucleolin in the nucleus, and mediate translocation of a glucose transporter between internal and plasma membranes (9 -16). C. elegans PKC3 is required for polarized accumulation of several regulatory proteins along portions of the periphery of 1-cell embryos (17, 18). Thus, aPKCs apparently regulate effector proteins at multiple intracellular locations.Mechanisms by which aPKCs are activated and targeted to specific microenvironments are poorly understood. All PKCs have C-terminal catalytic domains and N-terminal pseudosubstrate sites and Cys-rich regions (C1 domains) (1, 2, 4); phosphatidylserine stimulates catalytic activity of all PKCs. However, aPKCs do not bind Ca 2ϩ , DAG, or phorbol esters (which mimic DAG) and are not activated or translocated by these molecules (19 -21). aPKCs also lack transmembrane domains, cytoskeleton attachment sites, and organelle targeting motifs. Unlike other PKC isoforms, aPKCs escape endosome-mediated degradation when cells are incubated chronically with hormones, DAG analogs, or phorbol esters (19 -21). Thus, the paradigm of DAG-mediated membrane translocation/activation and subsequent degradation cannot explain targeting and regulation of aPKCs.Recent evidence suggests a "recruitment model" for incorporation of aPKCs into signaling pathways. aPKCs exhibit sub-* This wo...

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Structure of a Numb PTB domain–peptide complex suggests a basis for diverse binding specificity

Cited by 111 publications

References 48 publications

Autoinhibition of Mint1 adaptor protein regulates amyloid precursor protein binding and processing

Autoinhibition of Mint1 adaptor protein regulates amyloid precursor protein binding and processing

Molecular Basis for Interaction between Icap1α PTB Domain and β1 Integrin

A Novel Adapter Protein Employs a Phosphotyrosine Binding Domain and Exceptionally Basic N-terminal Domains to Capture and Localize an Atypical Protein Kinase C

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