The Eukaryotic Linear Motif resource for
Functional Sites in Proteins
Functional site class:
GTPase-binding domain (GBD) ligand
Functional site description:
The C helix is an amphipatic alpha helix located in the C-terminal VCA segment of WASP and N-WASP proteins. This helix binds to the centrally located GBD region of the same protein. This intramoleular interaction hides the acidic domain in the VCA segment from the Arp2/3 complex, thus preventing Arp2/3-dependent activation of actin polymerization. Some pathogens use a mimic of this motif in a multivalent manner to hijack cell regulation and promote actin polymerisation.
ELM Description:
The alpha helix has at least three turns where the positions +1, +5 and +9 need to be hydrophobic as they face a hydrophobic cavity in the GBD region of either WASP or N-WASP (Panchal,2003; Sallee,2008; Cheng,2008; Okrut,2015). The second position is also occupied by a hydrophobic residue, Val in the case of WASP, N-WASP, NCK1 and NCK2, and Ala in the two bacterial proteins.

Positions +3 and +7 are completely oriented away from the GBD region, therefore no restriction exists for these two positions.

Position +4 does interact with the GBD region, showing an Ala in WASP, N-WASP and the bacterial EspF. Based on the structure of the autoinhibited WASP, this Ala interacts with the Phe 293 in WASP. In the case of Nck1, an Asn is present while EspFU has an Arg that also interacts with Phe293. This indicates that the +4 position is variable but influences the binding with the target domain.

Position +6 has a conserved Met in WASP and N-WASP, a Lys in Nck1, Nck2 and EspF, and different nonpolar residues in the different repeats and homologs of EspFU (Met being the most common).

The structure of the autoinhibited WASP indicates that position +8 can only host a small amino acid like Val that could be interacting with Leu289, while the structure of EspFU bound to the GBD region in WASP shows that the corresponding residue in this position is slightly oriented outside the contacting area but still interacting with Asp92.

Dreadlocks (or Dock), the Drosophila homolog of NCK1, as well as other arthropod homologs, contains a Phe in the +2 position and different hydrophobic residues in the +9 position (Iso, Val). Dreadlocks is known to interact with WASp, the Drosophila homolog of WASP; however, the GBD-binding activity of the Phe-containing motif remains to be shown (Kaipa,2013).
Pattern: [ILV][VA][^P][^P][LI][^P][^P][^P][LM]
Pattern Probability: 0.0000979
Present in taxon: Chordata
Interaction Domain:
PBD (PF00786) P21-Rho-binding domain (Stochiometry: 1 : 1)
o See 12 Instances for LIG_GBD_Chelix_1
o Abstract
WASP and N-WASP proteins are required to initiate actin nucleation by activating actin-related protein 2/3 (Arp2/3) complex. WASP/N-WASP contains a GTPase-binding domain (GBD) at the N-terminus and a verprolin-homology, connector-helix, acidic motif (VCA) segment (also referred to as WCA as the V region is also called WH2) at the C-terminus. Under basal conditions the C-helix motif fits in the GBD domain, closing the protein and preventing its nucleation-promoting function (Kim,2000). Three factors participate in coordination to activate N-WASP at the membrane: (i) a GTPase like CDC42 that binds the GBD domain, (ii) an acidic phospholipid like PtdIns(4,5)P2, and (iii) an SH3-domain-containing protein (like Nck) that binds to the PxxP motifs located between the GBD and the VCA segment (Abdul-Manan,1999; Rohatgi,2001; Okrut,2015). Nck localises to the membrane using its SH2 domain to bind to phosphorylated tyrosine-containing proteins like Nephrin (ELMI002128), which is a transmembrane protein. The co-localization of Nck and N-WASP allows the C-helix motif of Nck, located in the linker sequence between the first and the second SH3 domain, to bind to the N-WASP GBD domain, thus outcompeting the intramolecular C-helix in N-WASP. Additional binding between N-WASP and Nck can occur due to interactions between the PxxP motifs in N-WASP and the second and third SH3 domain in Nck (Okrut,2015).

Interestingly, the effector proteins EspF and EspFU from the human pathogens enterohaemorrhagic and enteropathogenic Escherichia coli are able to activate WASP/N-WASP by using a mimic of the C-helix motif that is present in multiple copies, three in the case of EspF and between five and seven for EspFU (Alto,2007; Cheng,2008; Sallee,2008). The motif is conserved in homologous proteins from Citrobacter rodentium, a mouse enteric bacterium. The actin polymerization activating potency is higher in the pathogenic proteins due to multivalent binding of the repeats, increasing the density of activated N-WASP molecules (Sallee,2008).
o 10 selected references:

o 10 GO-Terms:

o 12 Instances for LIG_GBD_Chelix_1
(click table headers for sorting; Notes column: =Number of Switches, =Number of Interactions)
Acc., Gene-, NameStartEndSubsequenceLogic#Ev.OrganismNotes
P16333 NCK1
65 75 SARKASIVKNLKDTLGIGKV TP 1 Homo sapiens (Human)
C6UYI3 espF(U)
177 185 RPLPDVAQRLVQHLAEHGIQ TP 3 Escherichia coli O157:H7 str. TW14359
C6UYI3 espF(U)
130 138 RPLPDVAQRLVQHLAEHGIQ TP 3 Escherichia coli O157:H7 str. TW14359
C6UYI3 espF(U)
318 326 RPLPDVAQRLMQHLAEHGIN TP 3 Escherichia coli O157:H7 str. TW14359
C6UYI3 espF(U)
224 232 RPLPDVAQRLMQHLAEHGIQ TP 6 Escherichia coli O157:H7 str. TW14359
C6UYI3 espF(U)
271 279 RPLPDVAQRLMQHLAEHGIQ TP 6 Escherichia coli O157:H7 str. TW14359
B7UM88 espF
189 197 GGLPPLAQALKDHLAAYEQS TP 2 Escherichia coli O127:H6 str. E2348/69
B7UM88 espF
143 151 RPLPPIAQALKDHLAAYELS TP 2 Escherichia coli O127:H6 str. E2348/69
B7UM88 espF
96 104 RPLPPIAQALKDHLAAYELS TP 2 Escherichia coli O127:H6 str. E2348/69
O00401 WASL
467 475 APTSGIVGALMEVMQKRSKA TP 4 Homo sapiens (Human)
O08816 Wasl
463 471 APTSGIVGALMEVMQKRSKA TP 1 Rattus norvegicus (Norway rat)
P42768 WAS
466 474 QSSEGLVGALMHVMQKRSRA TP 3 Homo sapiens (Human)
Please cite: The Eukaryotic Linear Motif resource: 2022 release. (PMID:34718738)

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