Protein Engineering Flashcards
(14 cards)
important bits of the definition
-modification of a protein sequence, or making a new peotein
-making a protein more suitable for a specific purpose
tool for generating a recombinant protein
site-directed mutagenesis, creates a pool which can be screened
3 methods of protein engineering
rational mutagenesis, directed evolution, de novo design
examples of rational mutagenesis
- RuBisCO- trying to alter land plant versions to align more with red algae versions, which are more efficient
-hypertrophic cardiomyopathy- single point mutation can improve the phenotype (also being developed for CF)
-developing PETases- thermostability mutations, introducing mutations to improve temp tolerance (also applies to a lot of other industrial stuff)
how can diversification be done for directed evolution
chemical mutagenesis, e.g. UV ratiation etc
non-chemical mutagenesis, e.g. deactivating proofreading enzymes
mutating targeted sites, synthetic DNA molecules
recombination
how can screening be done in directed evo
organismal survival- useful for AMR stuff
fluorescent readout
selections for binding affinity, e.g. using antibodies
directed evolution examples
-improving phage thermostability- mutating, and adding selection pressure (making it very warm)
-RuBisCO again- using a toxic reaction where the product can only be broken down by RuBisCO as the pressure
-can also be used to optimise temp/pH sensitivity
what is PACE
phage assisted directed evo- coupling phage infection with cell properties
coupling the protein of interest with the protein which allows the phage to target E coli and therefore replicate- e.g. evolving a protein to recognise a promoter for this gene
synthetic biology definition
construction of new biological parts, or modifying existing biological systems (e.g. by adding unnatural amino acids)
current alphafold tech
af3- recent- moels using individual atoms, rather than broad properties of AAs-
issues w implementation
bacterial/yeast hosts- awks as these things are sometimes being moved into other organisms (plants or animals)
potential for mutation, mostly when dealing with pathogens/anything purposely going against selection pressure
non-native amino acids- reducing translation efficiency for example
example of synthetic biology
-alphafold to make antivenoms
-attempts to attenuate viruses for vaccination via introducing unnatural amino acids
