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|Functional site class:||UEV Domain binding PTAP motif|
|Functional site description:||The PTAP motif mediates binding of several cellular and viral proteins to the UEV domain of the class E vacuolar sorting protein Tsg101. Like several other linear motifs, including the CtBP binding motif in adenovirus and USP7 binding motif in EBV, PTAP was first discovered in a viral protein. Gottlinger et al. observed that disruption of the p6 region of HIV gag caused defective virus budding and further investigation pinpointed 4 residues, PTAP, vital to viral egress. The UEV domain of Tsg101 was not identified as the binding partner until ten years later. The solved structure of HIV gag-p6 PTAP motif bound to the UEV domain of Tsg101 showed that the motif formed one turn of a left-handed type II polyproline helix, similar to the ligands of WW and SH3 domains, while the flanking regions form extended conformations. The analysis showed that there are extensive interactions between the defined residues of the motif and the binding site, an observation consistent with the strict motif definition.|
|Description:|| The motif has an unusually strict definition, being fixed in all positions with the exception of the second position allowing a threonine or a serine. A randomized nonapeptide library was used for phage display and screened against the UEV domain returning 29 peptides of the consensus [AP][ST]AP allowing a non-canonical alanine at the first position (Schlundt et al., 2009). However no substitutions were tolerated in the AP region in a peptide that conferred binding. Peptide substitution analysis of the gag-p6 peptide PEPTAPPEE, confirmed that alanine was tolerated, however, proline was strongly preferred (Schlundt et al., 2009). Nevertheless, no functional instances are known to utilize this sub-optimal variant.
The UEV domain-bound motif forms one turn of a left-handed type II polyproline helix while the flanking regions form extended conformations. There are extensive intramolecular interactions between the defined residues of the motif. This observation may explain the strict motif definition (Pornillos et al., 2002). The most commonly seen variant of the peptide contains additional C-terminal prolines that have been suggested to increase affinity by stabilising the left-handed type II polyproline helix. A 9 residue peptide from the p6 chain of gag, PEPTAPPEE binds with the same affinity as full length p6, suggesting that at most the immediately flanking residues are necessary for Tsg101 binding of gag (Pornillos et al., 2002).
From the limited data available, viral proteins seem to have evolved a stronger binding affinity than host proteins. In an analysis of the TomL1 protein, HIV gag was shown to competitively out bind the UEV binding motif of TomL1 (Yanagida-Ishizaki et al., 2008). The gag-p6 motif binds with an affinity of Kd 27 micromolar, however no host motifs tested (Vps37b - 55 micromolar, SCAMP3 - 1975 micromolar and Tal-PSAP:62 micromolar and PTAP:344 micromolar) have bound with similar strength.
|Pattern:||.P[TS]AP. (Probability: 0.0001081)|
|Present in taxons:||Homo sapiens Metazoa|
PDB Structure: 1M4P
The creation of multivesicular bodies (MVBs) is dependent on class E vacuolar protein sorting (VPS) proteins, the majority of which are part of four ESCRT (Endosomal Sorting Complexes Required for Transport) complexes (For review see Raibourg et al.; 2009. Williams et al., 2007). These complexes are sequentially recruited to the membranes of endosomes as cargos are sorted and targeted (although this model has been challenged (Nickerson et al., 2007)). Tsg101 is a component of the ESCRT-1 complex, the second component of the VPS pathway (Previously ESCRT-I was the first ESCRT complex however recently HRS/STAM has been given the name ESCRT-0). Tsg101 recognises ubiquitinated cargoes via an ~145 aa N-terminal UEV domain and recruits the downstream ESCRT complexes. UEV domains are similar to E2 ubiquitin ligases, however, they have lost their ability to transfer ubiquitin due to the lack of a cysteine in the active site necessary to form thioester bonds with ubiquitin.
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