The Eukaryotic Linear Motif resource for
Functional Sites in Proteins
Accession:
Functional site class:
GTP-dependent BART-binding ligand
Functional site description:
ARF and ARF-like (ARL) subfamily proteins act as molecular signaling switches, cycling between GDP- and GTP-bound conformations and conducting different functions in each conformation.
A ligand motif present in the N-terminal helix of ARL2/3 ensures its GTP-dependent binding to a hydrophobic groove of the effector protein BART/ BARTL1 respectively (Lokaj,2015, Zhang,2009). Another group of ARL2/3 effectors, formed by PDE6d, HRG4/Unc119a and Unc119b, utilize a different binding mode, in which the N-terminal helix does not directly mediate the binding but may have additional functions. In complex with Unc119a, a protein responsible for the delivery of myristoylated cargo to the cilia, ARL3 allosterically displaces the cargo, with the N-terminal alpha-helix specifically influencing the release mechanism (Ismail,2012). In the GDP-bound form of ARF/ARL proteins, the N-terminal helix is anchored to a hydrophobic pocket on the surface of the protein (Hillig,2000).
ELM Description:
The motif is present in the N-terminal helix of the ARL2/3 proteins, which is then connected to a typical G-protein domain through a short loop that is thought to allow for the flexibility of the helix and its ability to adopt different conformations upon binding to various effectors or to the small GTPase itself.

BART and BARTL1 form similar binding domains, with two interfaces, both essential for the GTP-dependent interaction with ARL2/3 respectively. One of the interfaces encloses the N-terminal alpha-helix of ARL2/3 and the hydrophobic pocket formed by three alpha-helices in BART/ BARTL1. The binding is ensured by an extensive interaction between hydrophobic residues of the LLxILxx[LM] motif in ARL2/3 and several hydrophobic residues of BART/BARTL1. In the case of ARL2, there are several hydrogen bonds additionally formed at the interface.

The motif is well conserved, and the residues crucial for the interaction are identified mostly by biochemical analysis. Mutations of L3D, L4D and L7D in ARL2 each can disrupt its binding to BART; the L3A mutation decreases the binding affinit; and L4A, L7A mutations do not have significant effect on the affinity; the mutation I6R in ARL2 abolished its binding with BART (Zhang,2009). Similarly to the ARL2, L3D, L4D and L7D effects, mutations in ARL3 disrupt the binding to its effector BARTL1, as does the mutation L10D. Surprisingly, the I6R mutation did not cause loss in measured affinity (Lokaj,2015). This data suggests that residues L3, L4, I6, L7 and L10 play an important role in establishing the interaction between ARL2/3 and BART/BARTL1 respectively.

It was observed that the N-terminal helix of ARL3 is essential for the ciliary localization of the protein and partially for the myristoylated cargo release from UNC119a. Nevertheless, it was shown that the identity of the whole ARL3 N-terminal helix is indispensable (but not sufficient) for cargo release (Ismail,2012).
Pattern: ^..LL[^P]IL[^P][^P][LM]
Pattern Probability: 1.292e-09
Present in taxon: Metazoa
Interaction Domain:
ARL2_Bind_BART (PF11527) The ARF-like 2 binding protein BART (Stochiometry: 1 : 1)
o See 2 Instances for LIG_ARL_BART_1
o Abstract
Small G-proteins are involved in regulation of a variety of cellular processes and act as molecular switches cycling between GDP- and GTP-bound states. ARF subfamily proteins are mainly crucial to regulation of vesicular transport and undergo so-called beta-register shift between the GDP and the GTP conformation that promotes the release of an ARF-specific N-terminal myristoylated alpha-helix (Hanzal-Bayer,2002). The ARF-like (ARL) proteins are more heterogeneous than ARFs, and have more diverse functions, but nevertheless they share the same signaling mechanism: during the GDP-GTP exchange ARL2 and ARL3 undergo a beta-sheet register shift similar to that of the ARFs. In contrast with ARFs, the N-terminal region is non-myristoylated for ARL2, and acetylated for ARL3, but both form alpha-helices that can adopt different conformations in complexes with different effectors of the two small GTPases.

The GTP-dependent interaction between ARL2 and its binding partner BART (ARL2BP) involves two interfaces, one of which is mediated mostly by the switch I region of ARL2, and the other one is ensured by a well-conserved sequence motif LLxILxx[LM] for the N-terminal helix of ARL2. The helix binds the hydrophobic pocket of the BART protein, formed by several BART alpha helices, and the residues L3, L4, I6 and L7 of ARL2 make an extensive hydrophobic contact with several BART residues. Additionally, several hydrogen bounds are formed at this interaction interface (Zhang,2009, 3DOE).

A similar binding pattern is revealed for ARL3-GTP in complex with its effector CCDC104/CFAR36, also called BARTL1 for its structural homology with BART. The interaction again involves two areas, both essential for the binding. The first area interface is formed predominantly by the switch I and II regions of ARL3. In the second interface the N-terminal amphipathic helix of ARL3, containing LLxILxx[LM] motif, is completely buried in the hydrophobic groove of BARTL1. Interestingly, the N-terminal helix of ARL3 is essential not only for the interaction with BARTL1, but for the ciliary localization of the small GTPase itself (Lokaj,2015).

This binding mode sets BART and BARTL1 apart from other effectors of ARL2/3, as well as other effectors of the Ras superfamily proteins, mainly because it directly involves the N-terminal switch. In complex with PDEd, the N-terminal helix of ARL2-GTP protrudes on the surface and has no interaction to either ARL2 or its effector protein (Hanzal-Bayer,2002). Based on this, the N-terminal helix of ARL2 in complex with Unc119 is predicted to be exposed to the solvent as well. Strikingly, the structure ARL3-GppNHp in complex with Unc119a shows that, although the typical beta-sheet register shift also takes place upon the nucleotide exchange in ARL3, the hydrophobic pocket on the surface of the small GTPase remains free to be occupied by its N-terminal helix, which remains at least partially attached to the surface, the location resembling that of the GDP-bound conformation of ARL3 (1FZQ) as well as other ARF/ARL subfamily proteins. It was also shown that the presence of the ARL3 helix is indispensible (but not sufficient) for the myristoylated cargo release from Unc119a (Ismail,2012).

Structural and/or functional defects of some ARLs may drive ciliopathies, as several of this subfamily proteins were shown to be involved in ciliogenesis and cilia maintenance. Namely, Retinitis Pigmentosa is associated with mutations in ARL3 (22884633, Kuhnel,2006). In addition, ARL2 and ARL3 are known to regulate various microtubule-dependent processes, such as tubulin dynamics and folding (Veltel,2008). The complex ARL2-GTP-BART was reported to enter mitochondria and to bind to ANT-1, an inner mitochondrial transmembrane protein responsible for antiport of ADP/ATP (Sharer,2002). Recent studies show that BART is essential for nuclear retention of STAT3 protein, and its activity is enhanced by the presence of ARL2 (Muromoto,2008).
o 9 selected references:

o 13 GO-Terms:

o 2 Instances for LIG_ARL_BART_1
(click table headers for sorting; Notes column: =Number of Switches, =Number of Interactions)
Acc., Gene-, NameStartEndSubsequenceLogic#Ev.OrganismNotes
P36404 ARL2
ARL2_HUMAN
1 10 MGLLTILKKMKQKERELRLL TP 4 Homo sapiens (Human)
2 
Q9WUL7 Arl3
ARL3_MOUSE
1 10 MGLLSILRKLKSAPDQEVRI TP 4 Mus musculus (House mouse)
1 
Please cite: The eukaryotic linear motif resource - 2018 update. (PMID:29136216)

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