Intrinsically Disordered Proteins

4/12/2012
BiRDI, CTU, December 2012
Intrinsically disordered proteins (IDPs): a newly recognized class of proteins

Prof. Dr. Sonia Beeckmans
Research Unit Protein Chemistry Vrije Universiteit Brussel, Brussels, Belgium
Some historical discoveries:
Old paradigms in Biochemistry:

1 gene (DNA)
transcription
Discovery of the genetic code (1961-66): non-overlapping linear triplets of bases, called codons, lead to linear polymers of amino acids (proteins).
Discovery of mRNA: Franois Jacob, Jacques Monod George Beadle, (1961). Edward Tatum (1940).
1 mRNA
translation
First protein amino acid sequence (insulin) determined by Frederick Sanger (1953); First 3D-structure of proteins (haemoglobin/myoglobin) determined by John Kendrew and Max Perutz (1960), and first 3D-structure of an enzyme (lysozyme) by David Phillips (1965);
1 protein
folding
1 structure 1 function
Hypothesis: proteins unfold in denaturing conditions because of conformational changes that expose certain (mostly aliphatic) amino acid side chains to the solvent (hypothesis by Hsien Wu, 1929); thermodynamic hypothesis telling that the folded state corresponds to the global minimal free energy of the protein (Christian Anfinsen, 1960-ies).
4/12/2012
Old paradigms in Biochemistry:

1 gene (DNA)
transcription
The 11 principle turned out to be wrong, in all cases:

alternative splicing: introns/exons (Phillip Sharp, Richard Roberts, 1977) different potential transcriptioninitiation sites use of alternative start codons post-translational modifications (PTMs) structural diversity, e.g. because of PTMs different types of quaternary structure same protein may fold into different structures proteins exist that do not fold but yet have a function (IDPs: intrinsicallly disordered proteins) (discovery since 2000)
1 mRNA
translation
1 protein
folding
1 structure 1 function
IDPs: proteins that defy the structure function paradigm

Enzyme catalysis
structure function paradigm

Allosteric interactions Assisted protein folding Protein engineering 3D-structure analysis Protein misfolding and disease Proteomics Biotechnology Biomedicine
De novo protein biosynthesis
Discovery of IDPs
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The traditional view:

LOCK AND KEY MECHANISM
In the conventional view, a protein folds up immediately into a unique and stable 3D shape, the key (left). Its shape perfectly matches and allows it to bind its ligand(s), the lock (right).
Prof. Dr. Sonia Beeckmans Prof. Dr. Sonia Beeckmans
This is what we see for many enzymes, hormones, receptors, defense proteins (antibodies,...), transport proteins, etc...
Some examples
An enzyme firmly binds its substrate(s) and brings catalytic residues in optimal position
transition state
uncatalyzed reaction catalyzed reaction
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Hen egg white lysozyme
Gram+ bacteria
Hen egg white lysozyme

E35 and D52 are catalytic residues. It was demonstrated that E35 has an unusually high pK of 6 (instead of 4.3-4.5), which is due to its hydrophobic environment. N37, N44, Q57, D101 and R114 side chains, and F34, E35, Q57, N59, A107 and V109 backbone C=O or NH groups position the ligand in the active site cleft. W 62 and W 63 side chains help positioning the substrate in a correct way by stacking interactions with the sugar ring C. Catalysis proceeds via a covalently linked intermediate involving D52
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Serine proteases
The first step is the formation of an acyl-enzyme intermediate via a tetrahedral transition state. The second step is hydrolysis of the acyl-enzyme intermediate: the histidine residue activates a nucleophilic water molecule, and the reaction proceeds via a second transition state.
aspartate histidine serine triad
Serine proteases
ES complex
Stabilization of the tetrahedral transition state intermediate

The so called "oxyanion hole" formed by backbone NH groups stabilizes the negative charge of the oxygen atom during the transition state.
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An antibody firmly binds its antigen
IgG
An antibody firmly binds its antigen
Antibody conformation (Fab fragment) with Antibody conformation (same Fab no antigen bound. fragment) in the presence of antigen. Two residues in the heavy chain (blue) and one residue in the light chain (red) are shown for orientation.
The antibody binding site changes its conformation when antigen binds (induced fit). But the antibody may also induce conformational changes in the antigen. The binding cavity enlarged and several groups have altered positions.
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A lectin firmly binds its saccharide ligand

Legume seeds in general contain lectins (storage proteins/organizers of storage proteins?) that are often present in high amounts. Seeds from Pterocarpus angolensis (mukwa tree) contain high concentrations of such a lectin. From 1 kg peeled and defatted seeds, 21.5 grams of mukwa lectin can be purified by affinity chromatography on mannose-Sepharose. This lectin folds as a classical legume lectin type of structure (canonical dimer). It shows Mn/Ca-dependent saccharide binding. The monosaccharide specificity is mannose/glucose, but the lectin preferentially
loop A loop B (omega loop) loop C (metalbinding loop)
binds oligosaccharides.
mannose
loop D (specificity loop) loop E
D86 G106 N138 F132
A lectin firmly binds its saccharide ligand

disordered
prim +1
disordered
G (1,4)GN(1,2)M(1,6) G (1,4)GN(1,2)M(1,3)
M (1,4)GN(1,4)GN F (1,6)
+3
no H-bonds formed with the lectin
+2
no contact with the lectin
From binding studies:

the best binder is extension of mannoses with GN(1,4) abolishes binding extension with galactose residues diminishes binding further extension with sialic acid further diminishes but does not prevent binding removal of GN(1,2) highly reduces the affinity
M: mannose; GN: N-acetylglucosamine; G: galactose; F: fucose
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All these enzymes, defense molecules, lectins,... are globular, folded proteins.
They rely on their correct 3-dimensional structure for proper binding of their ligands and for their activity: the proper orientation of many amino acid residues in the ligand-binding sites is a prerequisite for their activity.
In order to understand these molecules, we need to get knowledge about their structure and their folding properties.
What determines the 3D structure of such globular proteins?
Amino acids are classified by their R-groups

We discriminate 5 main classes of R-groups:
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Weak interactions are crucial to macromolecular structure and function: 1. hydrogen bonds
H-bonds between backbone oxygen and amide hydrogens, but also involving amino acid side chains H-bonds between base pairs
Directionality!!
H-bonds are very important in determining the 3D-structure of biological macromolecules such as DNA and proteins
Weak interactions are crucial to macromolecular structure and function: 2. ionic interactions
Coulombs law describes the attraction and repulsion between two charges In a medium, the force is decreased and depends on the material of the medium: Q1*Q2 F=
Q1, Q2 charges on the two bodies r distance between them
In vacuum: Fvac =
Q1*Q2 r2
*r2
dielectric constant of the medium = Fvac / F > 1
(Water) = 80
Two electrical charges of opposite sign attract each other in water with 1/80 the force than in vacuum
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Weak interactions are crucial to macromolecular structure and function: 3. hydrophobic interactions
Non-polar molecules interfere with waterwater interactions: non-polar molecules tend to CLUSTER together in aqueous solutions, because they force energetically unfavorable changes in the structure of water. In e.g. mixtures water-benzene, waterhexane, ... two layers are formed. Hydrophobic compounds brought in water
water molecules in their vicinity are constrained in their possible orientations. They form highly ordered cage-like shells.
Driving force for folding of globular proteins!!
Loss in entropy: S < 0 And since G = H T*S
G > 0
thus unfavorable.
Weak interactions are crucial to macromolecular structure and function: 4. Vander Waals interactions
The non-covalent associations between electrically neutral molecules are known as van der Waals forces
E n e rg y (k c a l/m o le ).
2 1.5
E=
B r12
A r6
Van der Waals forces are a combination of three types of interactions: dipolar molecule dipolar molecule
Repulsion
0.5
Van der Waals distance

0 0 -0.5 1 2 3 4 5 6 7 8
Attraction
-1
dipolar molecule neutral molecule neutral molecule neutral molecule
r (angstrom)
Their energy varies with the distance between the molecules. A and B: constants that differ for the different types of interactions. In this example: interaction between two carbon atoms.
In a macromolecule, all atoms tend to be at VdW distance from their neighbours.
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Weak interactions are crucial to macromolecular structure and function: 5. - stacking interactions
aromatic AA side chains of F, Y and W, and also H, prefer interactions with interplane angles of around 90
Weak interactions are crucial to macromolecular structure and function:

Although the non-covalent interactions described, i.e. hydrogen bonds ionic interactions hydrophobic interactions van der Waals interactions - stacking interactions are individually weak, their cumulative effect can be very significant and determines the final 3D-structure of macromolecules such as globular proteins. The formation of each of these bonds contributes to a net decrease in free energy of the system: the cumulative effect of many small binding forces becomes enormous.
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The different levels of protein structure
(a) Primary structure amino acid sequence, including disulfide bonds (b) Secondary structure particularly stable arrangements of AA residues giving rise to recurring structural patterns (-H, -PS, -T): this involves short-range interactions (c) Tertiary structure describes all aspects of the 3D-folding of the polypeptide: this involves long-range interactions (d) Quaternary structure arrangement of polypeptide subunits in oligomeric proteins (e) Quinary structure association of different proteins to form multi-protein clusters
Protein secondary structure

Alpha-helix .....and beta-pleated sheet
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Supersecondary structures, also called motifs or folds

are particularly stable arrangements of several elements of secondary structure and connections between them
-hairpin
-motif
Immunoglobulin fold
Domain structure of polypeptide chains

Polypeptides with more than a few 100 AA often fold into two or several stable, globular units called
domains:
part of protein sequence and structure that can fold, evolve, function, and exist independently from the rest of the protein chain; each domain (about 25500AA) forms a certain 3D-structure and often is independently stable.
Often these domains will retain their correct 3D structure when isolated from the rest of the polypeptide chain. short linker peptide
Example: one subunit of GAPDH clearly is built of two distinct domains, an NAD+ binding domain (red), and a glyceraldehyde-3-P binding domain (green).
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Protein quaternary structure ...and quinary structure
Glyceraldehyde 3P dehydrogenase
ATCase
Phosphoglycerate kinase
GroeL/GroeS chaperonin
What determines folding of globular proteins?

Summarizing:
The amino acid sequence determines the 3D-structure (Anfinsens experiments with RNase!!)
Globular proteins can loose their native (i.e. properly folded) conformation, thereby
also loosing their biological activity: when denaturants are added into the solution (urea, organic molecules, detergents, ...) when the solution is either too acidic or too basic, or when the solution is heated (or frozen).
They denature proteins by changing the properties of the solvent

When the denaturing conditions are reversed,
polypeptides (may) fold rapidly by a stepwise process.

The folding pathway of a large polypeptide chain is complicated, and not all principles are fully understood yet.
Globular proteins tend to burry most of their hydrophobic residues inside the molecule (avoiding contact with water).
But: not all proteins fold to a defined 3D structure.
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Classical globular proteins Structure:

Structure:
the native protein has a welldefined 3D structure.
IDPs
Functions:
enzymes structural proteins folding machineries hormones receptors defense proteins (antibodies, ...) (membrane) transport proteins tags to act as signals ...
the native free protein is disordered, i.e. it exists as a dynamic ensemble of different structures; after binding to a partner, the IDP can become fully or partly folded (O, MG, PMG), these structures thereby becoming the native state.
Functions:
entropic chains (usually non-folders) molecular recognition molecules (usually folders); these IDPs can be hub proteins in protein networks
Estimated occurrence:
(proteins with disordered regions of >40 consecutive residues)
Bacteria: 733% Archaea: 937% Eukaryotes: 3663%
Protein folding process

in case of a small single-domain globular protein:
the polypeptide subjected to denaturing conditions in vitro behaves as an ensemble of many different structures (how many different structures will there be?
Are there some short distance interactions, which later may facilitate the folding process?)
The transition state is an ensemble of partially folded structures
Intermediates in the protein folding process
...and what about IDPs?...

From: LD Cabrita et al. (2010) Curr. Opin. Struct. Biol. 20, pp. 33-45
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From: VN Uversky & AK Dunker (2010) Biochim. Biophys. Acta 1804, pp 1231-64
Model structures of a 100-residues long polypeptide chain: from ordered protein structure (O), to molten globule-like structure (MG), to extended premolten globule-like structure (PMG), to unfolded protein (coil). Relative hydrodynamic volumes occupied by the same 100residues long polypeptide chain in the same four conformations.
N
Possible denaturation pathway for classically folded globular proteins Possible folding pathways for IDPs
D N
What about the physical properties of these structures?
O: Ordered globular protein.

Relatively rigid but with certain internal motions, and sometimes also conformational switches (short flexible hinge regions). All molecules have nearly the same structure; thus, a CD/fluorescence signal gives information about each molecule in the sample.
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MG: molten globule.

Still a compact structure (the hydrodynamic radius is not more than 15% bigger than in O, i.e. the volume of the structure did not increase more than 50% (volume of a sphere is: V = 4/3r3) Nearly all secondary structure is already present. Practically no tertiary structure is formed yet. Increased accessibility to proteases when compared to O. Increased accessibility to hydrophobic fluorescent probes (e.g. ANS) when compared to O.
PMG: pre-molten globule.

Is a bit less compact than the molten globule (the hydrodynamic radius is about 45% bigger than in O, i.e. the volume is about 3 times larger than the volume of O). It has no tertiary stucture. It has some (up to 50%), but not all secondary structure. It is non-globular (it is a squeezed and partially ordered form of a coil). It can effectively interact with the hydrophobic fluorescent probe ANS, but weaker than the MG (there are less hydrophobic clusters formed than in MG).
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Coil: random coil structure.

Is an extended structure with maximal dimensions (the hydrodynamic radius is about 2.3 times the radius in O, i.e. the volume is about 12 times larger than that of O). It is an ensemble of rapidly interchanging conformations (thus, a CD signal does not give information about each molecule in the sample, rather it is a mean of the different signals of each of the individual conformations).
Protein quartet model of protein functioning: different phase states of a protein
ORDERED MOLTEN GLOBULE

PRE-MOLTEN GLOBULE RANDOM

COIL
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The traditional view:
From: T Chouard (2011) Nature 471, pp 151-153

Example: tumor-suppressor protein p53 containing an ordered globular domain (brown) and disordered segments (colors) that help it to interact with hundreds of partners.
From: T Chouard (2011) Nature 471, pp 151-153
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IDPs form a newly recognized class of proteins They exist and properly function without a well-defined folded structure They are common in many proteomes They occur more frequently in more complex organisms They do not obey the rule according to which a well-defined 3D structure needs to be acquired, in which key residues that are important for binding or catalysis are properly positioned in space IDPs are said to be natively unfolded Some IDPs are disordered all over, some have only disordered domains or disordered regions (IDRs) They differ from globular proteins in two respects: due to the absence of tertiary interactions they are not globular, and they also have less secondary structure but a higher amount of coil conformations They have higher numbers of residues AGRKQSPE, and lower numbers of residues CWFYIVLN, i.e. they are characterized by a high net charge and low mean hydrophobicity (they are unable to form a hydrophobic core) Some IDPs remain disordered all the time, while others get ordered upon binding to their target (= induced folding)

IDPs often have essential functions They are commmonly found in signal transduction pathways, in cell-cycle regulation, gene expression and chaperone activity Advantages of structural disorder: increased speed of interaction high specificity coupled with low binding strength possibility to have more than one function (multiple binding partners) enabling larger interaction surfaces in the complex with partners Intrinsically Unstructured Proteins
(=IDPs)
From: P Tompa (2005) FEBS Lett. 579, pp 3346-54
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Functions of IDPs
Three broad categories:
molecular recognition macromol./macromolecule & macromol./ligand protein modification
y

entropic chains Proteins involved in regulation and signaling
From: K Dunker et al. (2002) Biochemistry 41, pp. 6574-82
Example: voltage-gated K+ ion channel is an entropic clock

Ion channels are present in the plasma membrane of all cells and provide movement of organic ions. Together with ion pumps, they regulate cytoplasmic ion concentrations and membrane potentials. Closed state before Charges within critical transmembrane depolarization membrane helices cause these helices to move relative to the membrane in response to changes in the membrane potential.
Inactivation of the channel
Open state after membrane depolarization (conformational changes in the channel)
Disordered chain of 60 residues

After membrane polarization, the cytoplasmic side of the pore opening gets a negative charge facilitating an interaction with the positively charged ball domain
From: VN Uversky & KA Dunker (2010) Biochim. Biophys. Acta 1804, pp. 1231-1264
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The availability of molecular recognition features and short linear peptide motifs within an unstructured segment enables fishing for and gathering of different partners.
IDPs as hub proteins:

Conformational flexibility facilitates access to enzymes that introduce or remove PTMs and to effectors that read the posttranslational code.
Conformational variability enables a nearly perfect moulding to fit the binding surfaces of very diverse partners.
Prof. Dr. Sonia Beeckmans Prof. Dr. Sonia Beeckmans
From: J Gsponer & MM Babu (2009) Progress Biophys. Mol. Biol. 99, pp. 94-103
Example: human protein p300 (histone acetyltransferase)

Protein regulating transcription via chromatin remodeling and transcription regulation, and important in the processes of cell proliferation and differentiation. It binds up to 400 partners. It consists of several domains and contains multiple PTM sites. Association with transcriptional activators (incomplete list):
(TAZ-1)
(KIX)
(TAZ-2) (HAT: histone acetyl transferase)
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Example: human protein p300 (histone acetyltransferase)

Acetylation: 2, 636, 977, 981, 1020, 1024, 1103, 1336, 1473, 1499, 1542, 1546, 1549, 1550, 1551, 1554, 1555, 1560, 1568, 1569, 1583, 1590, 1637, 1674 Phosphorylation: 89, 285, 885, 887, 1038, 1734, 1834, 1857, 1859 Methylation: 580, 604, 2142 Sumoylation: 1020
PTMs
Protein NP_001420 from UniProtKB/Swiss-Prot: a protein that is for 50% intrinsically disordered
335
418
1051
1159
1297
1517-1582 1662-1727 1834
2414
335418: TAZ-1 domain (CH1, cysteine-histidine-rich) 566646: KIX domain 10511159: bromo domain 11981278: TAZ domain (CH2) 13061608: KAT11 domain (histone acetylation) Disordered regions become 16681708: zinc finger domain folded after binding to structured targets, 17271806: TAZ-2 domain 20502096: IBiD domain while disordered targets become folded after
binding to structured regions of the protein.
IDRs are substrate of twice as many kinases as structured proteins the same is true for other PTMs
Small groups
phosphorylation methylation acetylation ADP-ribosylation
-PO42 (from ATP) -CH3 (from S-adenosylmethionine) CH3-CO-NAD

(from acetylcoenzyme A)
Proteins ubiquitination
sumoylation neddylation
Keep in mind: PTMs modify the local charge density and/or hydrophobicity, they affect the proteins structural and folding properties.
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Questions
Q: are the IDRs also disordered in vivo, or does the crowding environment inside cells lead to folding or partial folding? A: this may depend to a certain extent on the protein: -synuclein remains disordered inside E.coli, while FlgM appears to gain some structure.
Source
Prof. Dr. Sonia Beeckmans Human red blood cell E. coli Mitochondrion Brewers yeast Rabbit white muscle Rat red muscle Rat heart
Protein concentration (mg/ml)

250320 200 270560 114 220275 190240 200250 After D. Goodsell
Questions
Q: are the IDRs also disordered in vivo, or does the crowding environment inside cells lead to folding or partial folding? A: this may depend to a certain extent on the protein: -synuclein remains disordered inside E.coli, while FlgM appears to gain some structure. Q: are IDPs not easily degraded? A: in vivo they are apparently not more easily degraded. Q: are IDPs in vivo not captured by chaperones/chaperonins?
Hsp70
Hsp60
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Questions
Q: are the IDRs also disordered in vivo, or does the crowding environment inside cells lead to folding or partial folding? A: this may depend to a certain extent on the protein: -synuclein remains disordered inside E.coli, while FlgM appears to gain some structure. Q: are IDPs not easily degraded? A: in vivo they are apparently not easily degraded. Q: are IDPs in vivo not captured by chaperones/chaperonins? A: No! IDPs are essentially different from
unfolded or misfolded globular proteins.

Q: when IDPs fold, do we see first folding, or first binding?
Different types of ligands are known to induce folding of proteins

Carp parvalbumin folds upon binding of two Ca-ions.
Globular proteins
T4 fibritin folds upon assembly of three foldon domains.
The DNA-binding domain of brinker protein folds upon binding to DNA.

IDPs
Unfolded pKID protein folds upon binding to the folded KIX domain.
From: T Kiefhaber et al. (2012) Curr. Opin. Struct. Biol. 22, pp. 21-29
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Two opposite mechanisms may describe coupled folding and binding

Binding to the ligand occurs in a fully/partially unfolded state and folding occurs in the context of the complex.
Most probable mechanism
Little experimental evidence
A molecule from a low-populated folded state is selectively binding to the ligand.

From: T Kiefhaber et al. (2012) Curr. Opin. Struct. Biol. 22, pp. 21-29
Questions
Q: How do IDPs recognize and bind their targets? A: MoRFs (molecular recognition features) are short motifs (10-70 residues)
within large intrinsically disordered segments that promote specific protein-protein interactions. Upon binding, MoRFs undergo disorder-to-order transition.
Q: How do MoRFs look like? A: MoRFs are classified into

three subtypes according to their structure in the folded state: A. -MoRFs forming -helices, B. -MoRFs forming -strands, C. -MoRFs forming structures without a regular pattern of backbone hydrogen bonds; D. is an example of a complex MoRF.
MoRFs are in red; the partners are in green, while partner interfaces are shown as grey surfaces.
From: J Gsponer & MM Babu (2009) Progress Biophys. Mol. Biol. 99, pp. 94-103
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Example: bacterial RNA degradosome

A multi-enzyme cluster degrading mRNAs and also selectively processing other types of RNA
Structured region: RNA degrading activity Natively unstructured region with several microdomains of 15-40 aa; functions as scaffold for binding of different partners:
orthologs
paralogs
Enolase: glycolytic enzyme (converts 2-P-glycerate into PEP; function??) RNase R: 3-5 hydrolytic exoribonuclease Helicase RhlB: ATP-dependent (unwinds and translocates RNA substrates) PNPase: phosphorolytic exoribonuclease Rho: transcription termination factor
From: MJ Marcaida et al. (2006) Trends Biochem. Sci. 31, pp. 359-365 JAR Worrall et al. (2007) Biochem. Soc. Trans. 35, pp. 502-507
Example: E.coli RNA degradosome

Arginine-rich RNA-binding regions
Domain structure of the monomer
499
Predicted coiled-coil region
PNPase trimer
Tetramer; dimer of dimers (magenta/grey spheres: Mg2+/Zn2+)
Enolase dimer, with the recognition microdomain from the RNase E (red spheres: Mg2+)
From: MJ Marcaida et al. (2006) Trends Biochem. Sci. 31, pp. 359-365
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Example: E.coli RNA degradosome

Model based on crystallographic and biochemical data: a supramolecular machine of more than 4.1 mega-Da (larger than a ribosome), which is dedicated to RNA processing and turn-over. Up to eight enolase dimers Up to eight helicase monomers, each consisting of an N-terminal (pink) and a C-terminal (cyan) domain
Three tetrameric catalytic domains (12 subunits)

Long, unstructured tails with microdomains (MoRFs) for partner recognition
Coiled-coil interaction
Four PNPase trimers (12 subunits)

From: MJ Marcaida et al. (2006) Trends Biochem. Sci. 31, pp. 359-365
Prediction of protein IDPs/IDRs

Database for disordered proteins http://www.disprot.org/predictors.php
This website links to a whole series of Protein Disorder Predictors
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Prediction of protein IDPs/IDRs
645 IDPs 1,379 IDRs
Potential impact of disease mutations in ordered and disordered regions IDPs have been shown to be implicated in human
diseases such as cancer, diabetes, neurodegenerative and cardiovascular disorders.
What are predictions telling us?

20-30% (at least) of disease-related mutations are mapped in predicted protein disordered regions 20% of them are predicted to cause disorder-to-order (DO) transitions
Ordered proteins: Disordered proteins:
mainly involved in metabolism, biosynthesis, catalysis mainly involved in regulation/signaling
Mutations in IDPs are expected to affect proteinprotein/DNA/RNA/ligand interactions & PTMs,
assembly of macromolecular complexes
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Potential impact of disease mutations in ordered and disordered regions

Globular proteins
IDPs
From: V Vacic & LM Iakoucheva (2012) Mol. Biosyst. 8, pp. 27-32
In conclusion
7-33% 36-63% IDPs 9-37%

Proteins fulfilling essential functions
30

Intrinsically Disordered Proteins

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4/12/2012

BiRDI, CTU, December 2012

Intrinsically disordered proteins (IDPs): a newly recognized class of proteins

Some historical discoveries:

Old paradigms in Biochemistry:

Prof. Dr. Sonia Beeckmans

Old paradigms in Biochemistry:

The 11 principle turned out to be wrong, in all cases:

IDPs: proteins that defy the structure function paradigm

structure function paradigm

De novo protein biosynthesis

The traditional view:

Prof. Dr. Sonia Beeckmans Prof. Dr. Sonia Beeckmans

uncatalyzed reaction catalyzed reaction

Hen egg white lysozyme

Prof. Dr. Sonia Beeckmans

Hen egg white lysozyme

aspartate histidine serine triad

Prof. Dr. Sonia Beeckmans

Stabilization of the tetrahedral transition state intermediate

An antibody firmly binds its antigen

Prof. Dr. Sonia Beeckmans

An antibody firmly binds its antigen

Prof. Dr. Sonia Beeckmans

A lectin firmly binds its saccharide ligand

loop D (specificity loop) loop E

D86 G106 N138 F132

A lectin firmly binds its saccharide ligand

From binding studies:

What determines the 3D structure of such globular proteins?

Amino acids are classified by their R-groups

Prof. Dr. Sonia Beeckmans

Prof. Dr. Sonia Beeckmans

dielectric constant of the medium = Fvac / F > 1

Driving force for folding of globular proteins!!

Loss in entropy: S < 0 And since G = H T*S

Van der Waals distance

dipolar molecule neutral molecule neutral molecule neutral molecule

In a macromolecule, all atoms tend to be at VdW distance from their neighbours.

Prof. Dr. Sonia Beeckmans

Weak interactions are crucial to macromolecular structure and function:

Prof. Dr. Sonia Beeckmans

The different levels of protein structure

Prof. Dr. Sonia Beeckmans

Protein secondary structure

Prof. Dr. Sonia Beeckmans

Supersecondary structures, also called motifs or folds

Prof. Dr. Sonia Beeckmans

Domain structure of polypeptide chains

Protein quaternary structure ...and quinary structure

Prof. Dr. Sonia Beeckmans

What determines folding of globular proteins?

They denature proteins by changing the properties of the solvent

polypeptides (may) fold rapidly by a stepwise process.

But: not all proteins fold to a defined 3D structure.

Classical globular proteins Structure:

Prof. Dr. Sonia Beeckmans

Protein folding process

The transition state is an ensemble of partially folded structures

Intermediates in the protein folding process

Prof. Dr. Sonia Beeckmans

...and what about IDPs?...

Prof. Dr. Sonia Beeckmans