Structural characterization and computational analysis of <scp>PDZ</scp> domains in <i>Monosiga brevicollis</i>

Gao, Melody; Mackley, Iain G.P.; Mesbahi‐Vasey, Samaneh; Bamonte, Haley A.; Struyvenberg, Sarah; Landolt, Louisa; Pederson, Nick J.; Williams, Lucy; Bahl, Christopher D.; Brooks, Lionel; Amacher, J.F.

doi:10.1002/pro.3947

Cited by 9 publications

(28 citation statements)

References 104 publications

(288 reference statements)

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“…The A9UPE9 and DLG1-1 PDZ domains were expressed and purified following similar protocols, as previously described and in the Materials and Methods [ 48 , 54 ]. The reporter peptide used for both proteins was a fluoresceinated sequence matching the C-terminus of the HPV18 E6 oncoprotein (sequence: FITC-RLQRRRETQV), or F* -HPV18 E6.…”

Section: Resultsmentioning

confidence: 99%

“…Starting with the HPV18 E6 C-terminal sequence, we designed a number of peptides to test specific peptide positions, including HPV16 E6, HPV18 E6, S RET T V, S RET D V, RRET T V, and RRET D V, where underlined residues differ from HPV18 E6. As a control, we also tested human SNX27, SHANK1, and GIPC PDZ targets previously studied, including GIRK3 (sequence: ESESKV), BPIX (WDETNL), GAIP (QSSSEA), TYRP1 (PNQSVV), and B1AR (ASESKV), as well as previously identified mbSHANK targets, including A9V7Z4 (EDETAL), A9UP44 (QSESRL), and A9UXE1 (QDETAL) [ 48 , 54 ]. Example competition experiment binding curves are in Figure 6 , and all K I binding affinity values are reported in Table 3 .…”

Section: Resultsmentioning

confidence: 99%

“…For example, there are eight proteins in the M. brevicollis proteome that contain both PDZ and SH3 domains, a domain architecture that is only present in choanoflagellates based on BLAST searches, including A9UNP0_MONBE, A9UPE9_MONBE, A9UPI8_MONBE, A9UUC6_MONBE, A9UYE7_MONBE, A9V111_MONBE, A9V6P1_MONBE, and A9V7E4_MONBE. Notably, proteins that contain both PDZ and SH2 domains appear to be unique to organisms from the phylum Choanozoa as well, and there are five M. brevicollis proteins that contain these domains: A9V6T4_MONBE, A9V7X5_MONBE, A9VA09_MONBE, A9VB90_MONBE, and A9VC25_MONBE [ 48 ].…”

Section: Discussionmentioning

confidence: 99%

“…For example, choanoflagellates have extensive signaling networks and form a molecular architecture that resembles a multicellular state, called rosettes [ 51 , 52 , 53 ]. In our previous work, we concluded that there are 178 PDZ domains in M. brevicollis , with 1–20 PDZ domains in a single protein [ 48 ]. We previously used protein biochemistry and structural biology to characterize several PDZ domains from the M. brevicollis proteome, including those from mbDLG, mbGIPC, and mbSHANK1 ( Figure 1 ) [ 48 , 54 ].…”

Section: Introductionmentioning

confidence: 99%

“…In our previous work, we concluded that there are 178 PDZ domains in M. brevicollis , with 1–20 PDZ domains in a single protein [ 48 ]. We previously used protein biochemistry and structural biology to characterize several PDZ domains from the M. brevicollis proteome, including those from mbDLG, mbGIPC, and mbSHANK1 ( Figure 1 ) [ 48 , 54 ]. These PDZ domain-containing proteins all have clear homologues in the human proteome, and we found that selectivity determinants are largely conserved, despite ~750 million years of evolution between these organisms [ 48 ].…”

Section: Introductionmentioning

confidence: 99%

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

et al. 2021

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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.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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

et al. 2021

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Additive energetic contributions of multiple peptide positions determine the relative promiscuity of viral and human sequences for PDZ domain targets

et al. 2023

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Protein–protein interactions that involve recognition of short peptides are critical in cellular processes. Protein–peptide interaction surface areas are relatively small and shallow, and there are often overlapping specificities in families of peptide‐binding domains. Therefore, dissecting selectivity determinants can be challenging. PDZ domains are a family of peptide‐binding domains located in several intracellular signaling and trafficking pathways. These domains are also directly targeted by pathogens, and a hallmark of many oncogenic viral proteins is a PDZ‐binding motif. However, amidst sequences that target PDZ domains, there is a wide spectrum in relative promiscuity. For example, the viral HPV16 E6 oncoprotein recognizes over double the number of PDZ domain‐containing proteins as the cystic fibrosis transmembrane conductance regulator (CFTR) in the cell, despite similar PDZ targeting‐sequences and identical motif residues. Here, we determine binding affinities for PDZ domains known to bind either HPV16 E6 alone or both CFTR and HPV16 E6, using peptides matching WT and hybrid sequences. We also use energy minimization to model PDZ–peptide complexes and use sequence analyses to investigate this difference. We find that while the majority of single mutations had marginal effects on overall affinity, the additive effect on the free energy of binding accurately describes the selectivity observed. Taken together, our results describe how complex and differing PDZ interactomes can be programmed in the cell.

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Mint/X11 PDZ domains from non-bilaterian animals recognize and bind CaV2 calcium channel C-termini in vitro

Harracksingh,

Singh,

Mayorova

et al. 2024

Sci Rep

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PDZ domain mediated interactions with voltage-gated calcium (CaV) channel C-termini play important roles in localizing membrane Ca2+ signaling. The first such interaction was described between the scaffolding protein Mint-1 and CaV2.2 in mammals. In this study, we show through various in silico analyses that Mint is an animal-specific gene with a highly divergent N-terminus but a strongly conserved C-terminus comprised of a phosphotyrosine binding domain, two tandem PDZ domains (PDZ-1 and PDZ-2), and a C-terminal auto-inhibitory element that binds and inhibits PDZ-1. In addition to CaV2 chanels, most genes that interact with Mint are also deeply conserved including amyloid precursor proteins, presenilins, neurexin, and CASK and Veli which form a tripartite complex with Mint in bilaterians. Through yeast and bacterial 2-hybrid experiments, we show that Mint and CaV2 channels from cnidarians and placozoans interact in vitro, and in situ hybridization revealed co-expression in dissociated neurons from the cnidarian Nematostella vectensis. Unexpectedly, the Mint orthologue from the ctenophore Hormiphora californiensis strongly bound the divergent C-terminal ligands of cnidarian and placozoan CaV2 channels, despite neither the ctenophore Mint, nor the placozoan and cnidarian orthologues, binding the ctenophore CaV2 channel C-terminus. Altogether, our analyses suggest that the capacity of Mint to bind CaV2 channels predates bilaterian animals, and that evolutionary changes in CaV2 channel C-terminal sequences resulted in altered binding modalities with Mint.

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Structural characterization and computational analysis of PDZ domains in Monosiga brevicollis

Cited by 9 publications

References 104 publications

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

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

Additive energetic contributions of multiple peptide positions determine the relative promiscuity of viral and human sequences for PDZ domain targets

Mint/X11 PDZ domains from non-bilaterian animals recognize and bind CaV2 calcium channel C-termini in vitro

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