Short linear motif

The Human papilloma virus E7 oncoprotein mimic of the LxCxE motif (red) bound to the host Retinoblastoma protein (dark grey)(PDB: 1gux​)

In molecular biology Short Linear Motifs (also known as SLiMs, Linear Motifs or minimotifs) are short stretches of protein sequence that mediate protein protein interaction.

The first definition was given by Tim Hunt:

“The sequences of many proteins contain short, conserved motifs that are involved in recognition and targeting activities, often separate from other functional properties of the molecule in which they occur. These motifs are linear, in the sense that three-dimensional organization is not required to bring distant segments of the molecule together to make the recognizable unit. The conservation of these motifs varies: some are highly conserved while others, for example, allow substitutions that retain only a certain pattern of charge across the motif.”

Attributes

SLiMs are usually encoded in less than 10 contiguous amino acid residues, however, often only 2-4 of these residues contribute the majority of their binding specificity and affinity. Due to the limited contacts with their partner globular binding domain, SLiM mediated interactions are generally of low affinity, usually in the micromolar range. As a result, SLiM interactions are often transient and easily reversible, rendering them ideal for highly dynamic processes such as cell signalling. However, co-operative use can allow multiple low affinity motif create high avidity interactions, equivalent to domain-domain interfaces. For example, some endocytic adapter protein (e.g. in Eps15 protein) contain at least 15 repeats of an Asp-Pro-Phe tripeptide, which are responsible for interaction with multiple copies of the AP2 alpha appendage protein.

As a result of the limited number of critical amino acid positions in a SLiM, they can easily convergently evolve, as only a few mutations are necessary to create a rudimentary instances of a motif. Affinity and specificity can then be tuned by further mutations in the flanking residues.

They are typically (around 85% of occurrences) found in unstructured disordered regions of proteins. However, binding to a partner domain often induces SLiMs to form secondary structure, usually binding as an alpha helix or as a beta strand by beta augmentation.

Function

There are four main classes of linear motif:

1. Protein binding motifs bind the domain of interacting proteins and are the most common linear motifs. They may for example be involved in co-operative assembly of scaffolds. A typical example of such motifs are proline-rich sequences that are responsible for binding of SH3 domains. The class of proteins containing these motifs are usually ligands of the domain that binds them.
2. Some linear motifs describe regions of the protein recognised by post-translational modification enzymes such as sites of N-linked glycosylation.
3. A class of linear motifs are those involved in targeting proteins to different subcellular locations such as the C-terminal KDEL motif which marks proteins for endoplasmic retention.
4. Finally, some linear motifs are peptidase cleavage sites which are sites of proteins that are cut by enzymes. The motifs in most cases represent a segment of the protein that interacts with another protein.

Role in disease

Several diseases have been linked to mutations in SLiMs. For instance, one cause of Noonan Syndrome is a mutation in the protein Raf-1 which abrogates the interaction with 14-3-3 proteins mediated by corresponding short linear motifs and thereby deregulate the Raf-1 kinase activity . Usher's Syndrome is the most frequent cause of hereditary deaf-blindness in humans and can be caused by mutations in either PDZ domains in Harmonin or the corresponding PDZ interaction motifs in the SANS protein . Finally, Liddle's Syndrome has been implicated with autosomal dominant activating mutations in the WW interaction motif in the β-(SCNNB_HUMA) and γ-(SCNNG_HUMA) subunits of the Epithelial_sodium_channel ENaC . These mutations abrogate the binding to the ubiquitin ligase NEDD4, thereby inhibiting channel degradation and prolonging the half-life of ENaC, ultimately resulting in increased Na⁺ reabsorption, plasma volume extension and hypertension .

In addition, viruses mimic host motifs extensively to hijack and disrupt host protein functionality. The KDEL motif of the bacteriophage encoded cholera toxin mediates cell entry.

MDM2 SWIB domain-binding motif mimic drug Nutlin bound to MDM2(PDB: 3lbk​)

Potential as leads for drug design

Several motifs have been investigated as leads to design therapeutics such as the Integrin binding RGD-motif mimetic Cilengitide and the MDM2 SWIB domain-binding motif mimic drug Nutlin.

Computational motif resources

Databases

SLiMs are usually described by regular expressions in the motif literature with the important residues defined based on a combination of experimental, structural and evolutionary evidence. However, high throughput screening such as phage display has seen a large increase in the available information for many motifs classes allowing them to be described with sequence logos. Several diverse repositories currently curate the available motif data. In terms of scope, the Eukaryotic Linear Motif resource (ELM) and MiniMotif Miner (MnM) represent the two largest motif databases as they attempt to capture all motifs from the available literature. Several more specific and specialised databases also exist, PepCyber and ScanSite focus on smaller subsets of motifs, phosphopeptide binding and important signaling domains respectively. PDZBase focuses solely on PDZ domain ligands. Merops and CutDB curate available proteolytic event data including protease specificity and cleavage sites.

There has been a large increase in the number of publications describing motif mediated interactions over past decade and as a result a large amount of the available literature remains to be curated. Recent work has created the tool MiMosa to expedite the annotation process and encourage semantically robust motif descriptions.

Discovery tools

SLiMs are short and degenerate, and as a result the proteome is littered with stochastically occurring peptides that resemble functional motifs. The biologically relevant cellular partners can easily distinguish functional motifs, however computational tools have yet to reach a level of sophistication where motif discovery can be accomplished with high success rates.

Several attributes of motifs allow obvious false positives to be discriminated and the most likely true positives to selected/ranked.

• Accessibility
• Conservation
• Physicochemical properties
• Enrichment in groupings of similar proteins

Motif discovery tools can be split into two major categories, discovery of novel instance of known functional motifs class and discovery of functional motifs class.

Novel functional motifs instances

The Eukaryotic Linear Motif resource (ELM) and MiniMotif Miner (MnM) both provide servers to search for novel instance of known functional motifs in protein sequences. SLiMSearch allows similar searches on a proteome-wide scale .

Novel functional motifs class

More recently computational methods have been developed that can identify new Short Linear Motifs de novo. Interactome-based tools rely on identifying a set of proteins that are likely to share a common function, such as binding the same protein or being cleaved by the same peptidase. Two examples of such software are DILIMOT and SLiMFinder.. Anchor and α-MoRF-Pred use physicochemical properties to search for motif-like peptides in disordered regions. ANCHOR identifies stretches of intrinsically disordered regions that cannot form favorable intrachain interactions to fold without additional stabilising energy contributed by an interaction partner globular protein partner. α-MoRF-Pred inherent propensity of many SLiM to under go a disorder to order transition upon binding to discover α-helical forming stretches within disordered regions.

External links

• Pawsons Lab Resource on motif-binding domains

SLiM databases

• Eukaryotic Linear Motif Database
• MiniMotif Miner
• PepCyber
• ScanSite

SLiM discovery tools

• ANCHOR
• Eukaryotic Linear Motif Database
• MiniMotif Miner
• SLiMSuite : Website for SLiMFinder, SLiMSearch and CompariMotif
• ScanSite