Lecture 20 (Dobbek) Flashcards
Protein Folding and Evolution
Evolutionary Cycle
Evolutionary Cycle
- Selection acts on the Protein Function
Levels at which proteins evolve
Levels at which proteins evolve
Individual domains
- Minor changes in the overall structure (e.g. Amino acid substitutions, deletions and insertions)
Higher Structural Level
- Recombining existing domains to form new proteins
Challenges in the evolution of proteins
Challenges in the evolution of proteins
- We need reliable methods to detect homologous proteins
- Understand the allowable evolutionary pathways connecting homologous proteins
- Understand what constrains the divergence of proteins sequences, structures and functions (Function might be lost)
Homology: descendence from a common ancestor.
–> Inferring homology from similarity in sequence, structure and function (indirect because LUCA doesn’t excist anymore)
- Calibration of our methods to provide a threshold for confident conclusions about homology (because similarities don’t always mean homology)
- To link distant homologs with confidence the inclusion of intermediate proteins can be advantagous (to get idea of what happened during evolution)
- Distant relatives are best recognizable by their similar structures/folds (better than sequence comparison)
How can new proteins evolve?
How can new proteins evolve?
Gene Duplications
- Gene either gets lost (e.g. through stop codon mutation) due to selective pressure on one gene only
- Genes remain similar due to selective pressure on both genes
- One Copy aqquires a new function (selective advantage) –> most unlikely
- Gene duplications allow to acquire new functions, and consequently new proteins
Orthologs and Paralogs
Orthologs and Paralogs
- Homolog: Genes/entities that descend from a common ancestor
- Ortholog: if we can trace back to the last common ancestor (LCA) of the two genes without a gene duplication event AND if the LCA gave rise to the genes via population lineage-splitting event
- Paralog: if we can trace back to the last common ancestor (LCA) of the two genes including a gene duplication event
Exampe of Orthologs and Paralogs:
Hemoglobin
Exampe of Orthologs and Paralogs: Hemoglobin
- Gene duplication of the Monomeric Hemoglobin created Hba and Hbß
Divergent Evolution
Divergent Evolution
- Speciation Events: New organism and in them are protein evolving
- We can connect the evolution of proteins to the evolution of animals
Convergent Evolution
Convergent Evolution
- Similarity due to finding a “solution” for the same “problem”
- e-g. Chymotrypsin and Subtilisin are both catalyzing the cleavage of a peptid-bond (evolved independently from another)
- e.g. Carboanhydrases (hydrate Carbodioxid) have the same active center (Zn and 3 His), but the rest of the structure is different due to independent evolution
Divergence and structural change
Divergence and structural change
- Core: Assembly of the central helices and sheets that usually retains its topology
- Peripheral regions: outside the core, may change their folding pattern completely
Concept of Protein Space
Concept of Protein Space
- Protein Space: all possible amino acid sequences arranged, so that two sequences are neighbors if one can be converted into the other by a single amino acid substitution.
- It follows that if evolution by natural selection is to occur, functional proteins must form a continuous network which can be traversed by unit mutational steps without passing through nonfunctional intermediates.
Evolution of the spatial architecture
Evolution of the spatial architecture
Two sources of genetic variations cause changes in protein structure
- Darwinian selection for functionally advantagous mutations
- Neutral evolution, whereby mutations are accumulating by
random drift
Amino acid substitutions are constrained by
- intramolecular / intermolecular interactions (satisfaction of Hbonding potential to polar side chains!) ⟶ because they hold the domains together
- Accessibility to waters/lipids
Protein Folding - Anfinsen experiment
Protein Folding – Anfinsen’s Experiment
- Proteins can denature due to mechanical, thermal, or chemical stress.
- The experiment aimed to study the relationship between a protein’s amino acid sequence, its three-dimensional conformation (fold), and its function.
- Anfinsen used ribonuclease, an enzyme that hydrolyzes ribonucleotides, and disrupted its 3D structure (including disulfide bonds) using urea and β-mercaptoethanol to determine the conditions required for restoring its structure and activity.
- The denatured ribonuclease lost its enzymatic activity, raising the question of whether the loss was due to chemical interference.
- After removing the denaturing chemicals, the protein refolded and regained its function, demonstrating that its amino acid sequence alone contained the necessary information for proper folding.
- However, when the protein refolded in an oxidized state immediately after denaturation, incorrect disulfide bonds formed, leading to improper folding.
- By adding traces of β-mercaptoethanol, these incorrect bonds were reduced and rearranged, allowing the protein to correctly fold and fully restore its function.
- This meant that the structure of the protein detemines function and the amino acid sequence alone determines the fold
Protein folding: Nucleation-Condensation
Protein folding: Nucleation-Condensation
- Levinthal paradox: Defined folding pathway by keeping partially correct intermediates
- Molten Globule: intermediate in protein folding with compact conformation and native-like secondary structure. Packing is typically less dense than in native state
Typical Folding Scenario
- Fast collapse to molten globule state
- Appearance of tertiary structure
- Formation of dense packing –> Native State
Protein folding: kinetics + thermodynamics
Protein folding: kinetics + thermodynamics
- Denatured to Native State is spontaneous (ΔG < 0) (energy available to do work)
- Proteins could be destabilized through the formation of a few H-bonds
- The contribution from Enthalpie (ΔH) (Possibilities of interactions from side chains) and Entropy (ΔS) (Entropy is negative) is different
