Flashcards in Protein engineering Deck (13)
What is a protein's secondary structure?
Higher-level structures formed from backbone interactions between amino acids
Describe α helices
Carbonyl of one amino acid and the amino group of an amino acid 4 down the chain hydrogen bond
Turn every ~3.6 amino acids
Side chains stick out
Describe β strands/sheets
Hydrogen bonding between carbonyl and amino group across parts of the primary structure
Can be antiparallel (planar H bonding and N- and C- termini are adjacent to partner) or parallel (Staggered H bonding where N- and C- termini is adjacent)
Describe disulfide bonds
Links between adjacent cysteine residues in oxidising conditions
More common in extracellular proteins and eukaryotes
What are structural motifs?
Arrangements of multiple secondary structure elements together
What are active sites?
Spatial organisation of catalytic residues and ligand binding pockets to give a protein its function
What are fusions/deletions?
Removal of residues and domains or the addition of tags
What is site-directed mutagenesis?
Codon indels by mismatch PCR
What is directed evolution?
Mimics natural evolutionary selection with specific libraries and selection for specific protein characteristics
What are the pros of directed evolution?
Doesn't require detailed structural knowledge of protein of interest
What are the cons of directed evolution?
PCR based methods can be biased
Selection method required
Needs high-throughput assay that measures what you want to evolve
What are some ways to generate a library?
Random libraries (buy)