Lecture 3: IDPs Flashcards
What are the characters of IDPs?
IDPs are biologically active but they do not adopt a well-defined globular fold.
• Lack of specific tertiary structure.
• Many have some residual secondary structure in some regions but lack them in an overall ordered manner.
• They may contain secondary structure (molten-globule or pre-molten-globule like).
• They populate an ensemble of conformations instead of a single one.
• Amino acid sequence is responsible.
• Unfolded due to hydrophobic effect. There is not enough of a large hydrophobic core to fold spontaneously. Lowe content of hydrophobic amino acids.
• Some of them are disordered throughout their full length.
• Some have extensive residues with disorder.
• Sequences have low hydrophobicity.
How do we detect and study them?
- There is no secondary structure shown from far-UV CD or FT-IR. There will be an increase in far UV CD ellipticity in a spectrum similar to random coil proteins.
- There is no tertiary structure shown from near-UV CD, fluorescence and poor dispersion in 1D and 2D NMR spectra.
- SAXS, gel filtration and dynamic light scattering shows expansion. They show different Stokes radii and radii or gyration.
- There will be missing electron density in X-ray structures and evidence of fast backbone dynamics from 15N NMR relaxation measurements.
- 1H-15N NMR HSQC spectra will be very different. There will be a lack of dispersion in the 1HN dimension (F2) due to a lack of secondary and tertiary structure.
- 1H, 13C and 15N NMR can give secondary structure information. 15N relaxation can tell us about backbone dynamics and flexibility.
What are the functions of IDPs?
- IDPs have a range of important functions. They are involved in cell signalling through cell cycle control and transcriptional and translational regulation.
- The advantage of IDPs is threefold: they can bind to a large surface area, they can bind with low affinity and high specificity and they can bind to several different ligands (they are often hub proteins in interaction networks).
- Rapid turnover of IDPs gives rapid regulatory control in response to changing environmental conditions.
What are effector IDPs?
Effectors are involved in regulation.
• P21 and P27 also have a role as inhibitors of CDKs. P27 is intrinsically disordered with partner proteins. It forms a complex with Cdk2 and cyclin A, while still preserving an open, extended conformation. There are multiple contact points over a large surface area for a relatively small protein.
• The GTPase-binding domain (GBD) of WASP is disordered but it folds when it binds to its targets. WASP is autoinhibited by intramolecular interaction.
• When GBD binds to the VCA domain of WASP, the GBD folds into a compact globule with a hydrophobic core. In the presence of Cdc42 GTPase and Pip2, the GBD dissociates. A region GBD forms another strand of the Cdc42 beta sheet and a beta hairpin and a short helix are stabilised.
What are entropic chain activities?
Their functions stem from flexibility, pliability and plasticity of the backbone.
• For example, the titin PEVK domain gives enthalpic elasticity.
• Another use is the shaker-type voltage-gated potassium ion channel with a ball and chain timing mechanism.
• The timing of opening function depends directly on the length and flexibility of the disordered chain. A shorter chain causes a more rapid inactivation.
How do scavengers work?
Scavengers store or neutralise small ligands.
• For example, caseins bind calcium phosphate and inhibit ppt in milk.
How do assemblers work?
Display sites mediate regulatory post-translational modifications such as phosphorylation or limited proteolysis.
• For example, MAP2 microtubule binding domain interacts with protein kinases.
How do display sites work?
Display sites mediate regulatory post-translational modifications such as phosphorylation or limited proteolysis.
• For example, MAP2 microtubule binding domain interacts with protein kinases.
How do folding and binding work?
Does folding occur before binding or vice-versa? There are 2 mechanisms: induced folding or conformational selection.
• pKID of CREB is unstructured normally but it folds upon binding to KID-binding domain of CBP. The interactions overcome the entropic cost.
What are the genetics of IDPs?
The distribution of IDPs differs across species and domains.
• It is 4% for bacterial proteins.
• It is around 33% for eukaryotic proteins.
• It is thought that compartmentalisation allows disordered proteins to be protected from degradation. Prokaryotes also require increased selective pressure on biochemical efficiency.
• In mammals around 25-30% are mostly disordered. More than 50% of all proteins have regions of 30 or more residues. This is increased to more than 70% for signalling proteins.
• Disordered regions can be predicted and discovered. They tend to have lower overall hydrophobicity and a high overall net charge. There is a low sequence complexity. We predict disordered regions using these factors. PONDRs (predictors of natural disordered regions) use primary sequence data at an 80% accuracy.
• IDPs are also associated with RNA binding. Around 20% of RNA binding proteins were disordered to a significant extent.
