scholarly journals Domain Analysis and Motif Matcher (DAMM): A Program to Predict Selectivity Determinants in Monosiga brevicollis PDZ Domains Using Human PDZ Data

Molecules ◽  
2021 ◽  
Vol 26 (19) ◽  
pp. 6034
Author(s):  
Haley A. Wofford ◽  
Josh Myers-Dean ◽  
Brandon A. Vogel ◽  
Kevin Alexander Estrada Alamo ◽  
Frederick A. Longshore-Neate ◽  
...  
Keyword(s):  
Signaling Pathways ◽  
Pdz Domain ◽  
Peptide Binding ◽  
Domain Analysis ◽  
Cellular Adhesion ◽  
Pdz Domains ◽  
Cellular Targets ◽  
C Terminus ◽  
Domains Of Life ◽  
Monosiga Brevicollis

Choanoflagellates are single-celled eukaryotes with complex signaling pathways. They are considered the closest non-metazoan ancestors to mammals and other metazoans and form multicellular-like states called rosettes. The choanoflagellate Monosiga brevicollis contains over 150 PDZ domains, an important peptide-binding domain in all three domains of life (Archaea, Bacteria, and Eukarya). Therefore, an understanding of PDZ domain signaling pathways in choanoflagellates may provide insight into the origins of multicellularity. PDZ domains recognize the C-terminus of target proteins and regulate signaling and trafficking pathways, as well as cellular adhesion. Here, we developed a computational software suite, Domain Analysis and Motif Matcher (DAMM), that analyzes peptide-binding cleft sequence identity as compared with human PDZ domains and that can be used in combination with literature searches of known human PDZ-interacting sequences to predict target specificity in choanoflagellate PDZ domains. We used this program, protein biochemistry, fluorescence polarization, and structural analyses to characterize the specificity of A9UPE9_MONBE, a M. brevicollis PDZ domain-containing protein with no homology to any metazoan protein, finding that its PDZ domain is most similar to those of the DLG family. We then identified two endogenous sequences that bind A9UPE9 PDZ with <100 μM affinity, a value commonly considered the threshold for cellular PDZ–peptide interactions. Taken together, this approach can be used to predict cellular targets of previously uncharacterized PDZ domains in choanoflagellates and other organisms. Our data contribute to investigations into choanoflagellate signaling and how it informs metazoan evolution.

scholarly journals Domain Analysis and Motif Matcher (DAMM): A Program to Predict Selectivity Determinants in Monosiga brevicollis PDZ Domains Using Human PDZ Data

2021 ◽  
Author(s):  
Haley A. Wofford ◽  
Josh Myers-Dean ◽  
Brandon A. Vogel ◽  
Kevin Alexander Estrada Alamo ◽  
Frederick A. Longshore-Neate ◽  
...  
Keyword(s):  
Signaling Pathways ◽  
Pdz Domain ◽  
Peptide Binding ◽  
Domain Analysis ◽  
Cellular Adhesion ◽  
Pdz Domains ◽  
Cellular Targets ◽  
C Terminus ◽  
Domains Of Life ◽  
Monosiga Brevicollis

Choanoflagellates are single-celled eukaryotes with complex signaling pathways. They are considered the closest non-metazoan ancestors to mammals and other metazoans, and form multicellular-like states called rosettes. The choanoflagellate Monosiga brevicollis contains over 150 PDZ domains, an important peptide-binding domain in all three domains of life (Archaea, Bacteria, and Eukarya). Therefore, an understanding of PDZ domain signaling pathways in choanoflagellates may provide insight into the origins of multicellularity. PDZ domains recognize the C-terminus of target proteins and regulate signaling and trafficking pathways, as well as cellular adhesion. Here, we developed a computational program, Domain Analysis and Motif Matcher (DAMM), that predicts target specificity in choanoflagellate PDZ domains by analyzing peptide-binding cleft sequence identity as compared to human PDZ domains. We used this program, protein biochemistry, fluorescence polarization, and structural analyses to characterize the specificity of A9UPE9_MONBE, a M. brevicollis PDZ domain-containing protein with no homology to any metazoan protein, finding that its PDZ domain is most similar to those of the DLG family. We then identified two endogenous sequences that bind A9UPE9 PDZ with &amp;lt;100 M affinity, a value commonly considered the threshold for cellular PDZ-peptide interactions. Taken together, this approach can be used to predict cellular targets of previously uncharacterized PDZ domains in choanoflagellates and other organisms. Our data contributes to investigations into choanoflagellate signaling and how it informs metazoan evolution.

scholarly journals The evolution of non-motif selectivity determinants in Monosiga brevicollis PDZ domains

2020 ◽  
Author(s):  
Melody Gao ◽  
Iain G. P. Mackley ◽  
Samaneh Mesbahi-Vasey ◽  
Haley A. Bamonte ◽  
Sarah A. Struyvenberg ◽  
...  
Keyword(s):  
Signaling Pathways ◽  
Protein Interactions ◽  
Pdz Domain ◽  
Binding Motif ◽  
Binding Affinities ◽  
Pdz Domains ◽  
Protein Protein Interactions ◽  
Signaling Systems ◽  
Neuronal Synapses ◽  
Monosiga Brevicollis

AbstractThe evolution of signaling pathways is complex and well-studied. In particular, the emergence of animal multicellularity had a major impact on protein-protein interactions and signaling systems in eukaryotic cells. However, choanoflagellates, our closest non-metazoan ancestor, contain a number of closely related signaling and trafficking proteins and domains. In addition, because choanoflagellates can adopt a rosette-/multicellular-like state, a lot can be gained by comparing proteins involved in choanoflagellate and human signaling pathways. Here, we look at how selectivity determinants evolved in the PDZ domain. There are over 250 PDZ domains in the human proteome, which are involved in critical protein-protein interactions that result in large multimolecular complexes, e.g., in the postsynaptic density of neuronal synapses. Binding of C-terminal sequences by PDZ domains is often transient and recognition typically involves 6 residues or less, with only 2 residues specifying the binding motif. We solved high resolution crystal structures of Monosiga brevicollis PDZ domains homologous to human Dlg1 PDZ2, Dlg1 PDZ3, GIPC, and SHANK1 PDZ domains to investigate if the non-motif preferences are conserved, despite hundreds of millions of years of evolution. We also calculated binding affinities for GIPC, SHANK1, and SNX27 PDZ domains from M. brevicollis. Overall, we found that peptide selectivity is conserved between these two disparate organisms, with one exception, mbDLG-3. In addition, we identify 178 PDZ domains in the M. brevicollis proteome, including 11 new sequences, which we verified using Rosetta and homology modeling. Overall, our results provide novel insight into signaling pathways in the choanoflagellate organism.

scholarly journals Structures of a Human Papillomavirus (HPV) E6 Polypeptide Bound to MAGUK Proteins: Mechanisms of Targeting Tumor Suppressors by a High-Risk HPV Oncoprotein

2007 ◽  
Vol 81 (7) ◽  
pp. 3618-3626 ◽  
Author(s):  
Yi Zhang ◽  
Jhimli Dasgupta ◽  
Runlin Z. Ma ◽  
Lawrence Banks ◽  
Miranda Thomas ◽  
...  
Keyword(s):  
Human Papillomavirus ◽  
High Risk ◽  
Crystal Structures ◽  
Tumor Suppressors ◽  
Specific Binding ◽  
Pdz Domain ◽  
Pdz Domains ◽  
Hpv E6 ◽  
C Terminus ◽  
High Risk Hpv

ABSTRACT Human papillomavirus (HPV) E6 oncoprotein targets certain tumor suppressors such as MAGI-1 and SAP97/hDlg for degradation. A short peptide at the C terminus of E6 interacts specifically with the PDZ domains of these tumor suppressors, which is a property unique to high-risk HPVs that are associated with cervical cancer. The detailed recognition mechanisms between HPV E6 and PDZ proteins are unclear. To understand the specific binding of cellular PDZ substrates by HPV E6, we have solved the crystal structures of the complexes containing a peptide from HPV18 E6 bound to three PDZ domains from MAGI-1 and SAP97/Dlg. The complex crystal structures reveal novel features of PDZ peptide recognition that explain why high-risk HPV E6 can specifically target these cellular tumor suppressors for destruction. Moreover, a new peptide-binding loop on these PDZs is identified as interacting with the E6 peptide. Furthermore, we have identified an arginine residue, unique to high-risk HPV E6 but outside the canonical core PDZ recognition motif, that plays an important role in the binding of the PDZs of both MAGI-I and SAP97/Dlg, the mutation of which abolishes E6's ability to degrade the two proteins. Finally, we have identified a dimer form of MAGI-1 PDZ domain 1 in the cocrystal structure with E6 peptide, which may have functional relevance for MAGI-1 activity. In addition to its novel insights into the biochemistry of PDZ interactions, this study is important for understanding HPV-induced oncogenesis; this could provide a basis for developing antiviral and anticancer compounds.

scholarly journals Identification of PDZ Domain Containing Proteins Interacting with 1.2 and PMCA4b

2013 ◽  
Vol 2013 ◽  
pp. 1-16 ◽  
Author(s):  
Doreen Korb ◽  
Priscilla Y. Tng ◽  
Vladimir M. Milenkovic ◽  
Nadine Reichhart ◽  
Olaf Strauss ◽  
...  
Keyword(s):  
Pdz Domain ◽  
Pdz Domains ◽  
Hek 293 ◽  
Zonula Occludens ◽  
C Terminus ◽  
293 Cells ◽  
Interaction Partners ◽  
Voltage Dependent ◽  
Interaction Domains ◽  
Zonula Occludens 1

PDZ (PSD-95/Disc large/Zonula occludens-1) protein interaction domains bind to cytoplasmic protein C-termini of transmembrane proteins. In order to identify new interaction partners of the voltage-gated L-type Ca2+ channel 1.2 and the plasma membrane Ca2+ ATPase 4b (PMCA4b), we used PDZ domain arrays probing for 124 PDZ domains. We confirmed this by GST pull-downs and immunoprecipitations. In PDZ arrays, strongest interactions with 1.2 and PMCA4b were found for the PDZ domains of SAP-102, MAST-205, MAGI-1, MAGI-2, MAGI-3, and ZO-1. We observed binding of the 1.2 C-terminus to PDZ domains of NHERF1/2, Mint-2, and CASK. PMCA4b was observed to interact with Mint-2 and its known interactions with Chapsyn-110 and CASK were confirmed. Furthermore, we validated interaction of 1.2 and PMCA4b with NHERF1/2, CASK, MAST-205 and MAGI-3 via immunoprecipitation. We also verified the interaction of 1.2 and nNOS and hypothesized that nNOS overexpression might reduce Ca2+ influx through 1.2. To address this, we measured Ca2+ currents in HEK 293 cells co-expressing 1.2 and nNOS and observed reduced voltage-dependent 1.2 activation. Taken together, we conclude that 1.2 and PMCA4b bind promiscuously to various PDZ domains, and that our data provides the basis for further investigation of the physiological consequences of these interactions.

scholarly journals Steric clashes with bound OMP peptides activate the DegS stress-response protease

2015 ◽  
Vol 112 (11) ◽  
pp. 3326-3331 ◽  
Author(s):  
Anna K. de Regt ◽  
Tania A. Baker ◽  
Robert T. Sauer
Keyword(s):  
Outer Membrane Proteins ◽  
Pdz Domain ◽  
Peptide Binding ◽  
Structural Rearrangement ◽  
Signaling Cascade ◽  
Pdz Domains ◽  
Protease Domain ◽  
Allosteric Activation ◽  
Steric Clash ◽  
Envelope Stress

Escherichia coli senses envelope stress using a signaling cascade initiated when DegS cleaves a transmembrane inhibitor of a transcriptional activator for response genes. Each subunit of the DegS trimer contains a protease domain and a PDZ domain. During stress, unassembled outer-membrane proteins (OMPs) accumulate in the periplasm and their C-terminal peptides activate DegS by binding to its PDZ domains. In the absence of stress, autoinhibitory interactions, mediated by the L3 loop, stabilize inactive DegS, but it is not known how this autoinhibition is reversed during activation. Here, we show that OMP peptides initiate a steric clash between the PDZ domain and the L3 loop that results in a structural rearrangement of the loop and breaking of autoinhibitory interactions. Many different L3-loop sequences are compatible with activation but those that relieve the steric clash reduce OMP activation dramatically. Our results provide a compelling molecular mechanism for allosteric activation of DegS by OMP-peptide binding.

scholarly journals Requirement of PDZ-Containing Proteins for Cell Cycle Regulation and Differentiation in the Mouse Lens Epithelium

2003 ◽  
Vol 23 (24) ◽  
pp. 8970-8981 ◽  
Author(s):  
Minh M. Nguyen ◽  
Marie L. Nguyen ◽  
Georgina Caruana ◽  
Alan Bernstein ◽  
Paul F. Lambert ◽  
...  
Keyword(s):  
Pdz Domain ◽  
Cellular Adhesion ◽  
Pdz Domains ◽  
Mammalian Development ◽  
Normal Pattern ◽  
Viral Oncoprotein ◽  
Pdz Proteins ◽  
Cell Growth And Differentiation ◽  
Cycle Regulation ◽  
Mouse Lens

ABSTRACT The roles of PDZ domain-containing proteins such as Dlg and Scrib have been well described for Drosophila; however, their requirement for mammalian development is poorly understood. Here we show that Dlg, Scrib, MAGI1, MAGI3, and MPDZ are expressed in the mouse ocular lens. We demonstrate that the increase in proliferation and defects in cellular adhesion and differentiation observed in epithelia of lenses that express E6, a viral oncoprotein that can bind to several PDZ proteins, including the human homologs of Dlg and Scrib, is dependent on E6's ability to bind these proteins via their PDZ domains. Analyses of lenses from mice carrying an insertional mutation in Dlg (dlggt ) show increased proliferation and proliferation in spatially inappropriate regions of the lens, a phenotype similar to that of lenses expressing E6. The results from this study indicate that multiple PDZ domain-containing proteins, including Dlg and Scrib, may be required for maintaining the normal pattern of growth and differentiation in the lens. Furthermore, the phenotypic similarities among the Drosophila dlg mutant, the lenses of dlggt mice, and the lenses of E6 transgenic mice suggest that Dlg may have a conserved function in regulating epithelial cell growth and differentiation across species.

scholarly journals Interactomic affinity profiling by holdup assay: Acetylation and distal residues impact the PDZome-binding specificity of PTEN phosphatase

PLoS ONE ◽  
2020 ◽  
Vol 15 (12) ◽  
pp. e0244613
Author(s):  
Pau Jané ◽  
Gergő Gógl ◽  
Camille Kostmann ◽  
Goran Bich ◽  
Virginie Girault ◽  
...  
Keyword(s):  
Consensus Sequence ◽  
Pdz Domain ◽  
Pi3k Pathway ◽  
Binding Motif ◽  
Pdz Domains ◽  
Post Translational Modifications ◽  
Binding Profile ◽  
Protein Motifs ◽  
C Terminus ◽  
The Impact

Protein domains often recognize short linear protein motifs composed of a core conserved consensus sequence surrounded by less critical, modulatory positions. PTEN, a lipid phosphatase involved in phosphatidylinositol 3-kinase (PI3K) pathway, contains such a short motif located at the extreme C-terminus capable to recognize PDZ domains. It has been shown that the acetylation of this motif could modulate the interaction with several PDZ domains. Here we used an accurate experimental approach combining high-throughput holdup chromatographic assay and competitive fluorescence polarization technique to measure quantitative binding affinity profiles of the PDZ domain-binding motif (PBM) of PTEN. We substantially extended the previous knowledge towards the 266 known human PDZ domains, generating the full PDZome-binding profile of the PTEN PBM. We confirmed that inclusion of N-terminal flanking residues, acetylation or mutation of a lysine at a modulatory position significantly altered the PDZome-binding profile. A numerical specificity index is also introduced as an attempt to quantify the specificity of a given PBM over the complete PDZome. Our results highlight the impact of modulatory residues and post-translational modifications on PBM interactomes and their specificity.

scholarly journals Secondary PDZ domain-binding site on class B plexins enhances the affinity for PDZ–RhoGEF

2015 ◽  
Vol 112 (48) ◽  
pp. 14852-14857 ◽  
Author(s):  
Heath G. Pascoe ◽  
Stephen Gutowski ◽  
Hua Chen ◽  
Chad A. Brautigam ◽  
Zhe Chen ◽  
...  
Keyword(s):  
Pdz Domain ◽  
Specific Interaction ◽  
Cell Formation ◽  
General Mechanism ◽  
Nucleotide Exchange ◽  
Pdz Domains ◽  
Vast Number ◽  
C Terminus ◽  
Class B ◽  
Modular Domain

PDZ domains are abundant protein interaction modules and typically recognize a short motif at the C terminus of their ligands, with a few residues in the motif endowing the binding specificity. The sequence-based rules, however, cannot fully account for the specificity between the vast number of PDZ domains and ligands in the cell. Plexins are transmembrane receptors that regulate processes such as axon guidance and angiogenesis. Two related guanine nucleotide exchange factors (GEFs), PDZ–RhoGEF and leukemia-associated RhoGEF (LARG), use their PDZ domains to bind class B plexins and play critical roles in signaling. Here, we present the crystal structure of the full-length cytoplasmic region of PlexinB2 in complex with the PDZ domain of PDZ–RhoGEF. The structure reveals that, in addition to the canonical C-terminal motif/PDZ interaction, the 3D domain of PlexinB2 forms a secondary interface with the PDZ domain. Our biophysical and cell-based assays show that the secondary interface contributes to the specific interaction between plexin and PDZ–RhoGEF and to signaling by plexin in the cell. Formation of secondary interfaces may be a general mechanism for increasing affinity and specificity of modular domain-mediated interactions.

scholarly journals Channel-interacting PDZ protein, ‘CIPP’, interacts with proteins involved in cytoskeletal dynamics

2009 ◽  
Vol 419 (2) ◽  
pp. 289-300 ◽  
Author(s):  
Emanuele Alpi ◽  
Elena Landi ◽  
Manuela Barilari ◽  
Michela Serresi ◽  
Piero Salvadori ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Pdz Domain ◽  
Pdz Domains ◽  
Insulin Receptor Tyrosine Kinase ◽  
Kinase Substrate ◽  
Substrate Protein ◽  
C Terminus ◽  
Cytoskeletal Dynamics ◽  
Direct Binding ◽  
Pdz Protein

Neuronal CIPP (channel-interacting PDZ protein) is a multivalent PDZ protein that interacts with specific channels and receptors highly expressed in the brain. It is composed of four PDZ domains that behave as a scaffold to clusterize functionally connected proteins. In the present study, we selected a set of potential CIPP interactors that are involved directly or indirectly in mechanisms of cytoskeletal remodelling and membrane protrusion formation. For some of these, we first proved the direct binding to specific CIPP PDZ domains considered as autonomous elements, and then confirmed the interaction with the whole protein. In particular, the small G-protein effector IRSp53 (insulin receptor tyrosine kinase substrate protein p53) specifically interacts with the second PDZ domain of CIPP and, when co-transfected in cultured mammalian cells with a tagged full-length CIPP, it induces a marked reorganization of CIPP cytoplasmic localization. Large punctate structures are generated as a consequence of CIPP binding to the IRSp53 C-terminus. Analysis of the puncta nature, using various endocytic markers, revealed that they are not related to cytoplasmic vesicles, but rather represent multi-protein assemblies, where CIPP can tether other potential interactors.

Plasma membrane Ca2+-ATPase associates with CLP36, α-actinin and actin in human platelets

2007 ◽  
Vol 97 (04) ◽  
pp. 587-597 ◽  
Author(s):  
Larry Bozulic ◽  
Mohammad Malik ◽  
David Powell ◽  
Adrian Nanez ◽  
Andrew Link ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Pdz Domain ◽  
Gel Filtration ◽  
Human Platelets ◽  
Cytoskeletal Proteins ◽  
Pdz Domains ◽  
Glutathione S Transferase ◽  
Spectrometry Analysis ◽  
C Terminus ◽  
Domain Interactions

SummaryThe plasma membrane Ca2+-ATPase (PMCA) plays an essentialrole in maintaining low cytosolic Ca2+ in resting platelets. Earlier studies demonstrated that the 4b isoform of PMCA interacts viaits C-terminal end with the PDZ domains of membrane-associated guanylate kinase proteins. Activation of saponin-permeabilized platelets in the presence of a peptide composed of the lastten residues of the PMCA4b C-terminus leads to a significant decrease of PMCA associated with the cytoskeleton, suggesting that PDZ domain interactions play a role in tethering the pumpto the cytoskeleton. Here we present experiments conducted to evaluate the mechanism of this association. Co-immunoprecipitationassays coupled with liquid chromatography/tandemmass spectrometry analysis and immunoblotting were used to identify proteins that interact with PMCA in the resting platelet. Our results indicate that the only PDZ domain-containing proteinassociated with PMCA is the LIM family protein, CLP36. Glutathione-S-transferase pull-down from a platelet extractusing a fusion protein containing the C-terminal PDZ domainbinding motif of PMCA confirmed binding of CLP36 to PMCA. Gel filtration chromatography of detergent-solubilized plateletsdemonstrated the existence of a 1,000-kDa complex containingPMCA and CLP36, and in addition, α -actinin and actin. Immunoflourescencemicroscopy confirmed the co-localization ofPMCA with CLP36 in resting and activated platelets. Taken togetherthese results suggest that PMCA is localized in non-filamentousactin complexes in resting platelets by means of PDZdomain interactions and then associates with the actin cytoskeletonduring cytoskeletal rearrangement upon platelet activation. Thus, in addition to the reversible serine/threonine andtyrosine phosphorylation events previously described in humanplatelets, PMCA function may be regulated by interactions withanchoring and cytoskeletal proteins.

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