Lecture 16 Flashcards
What is conjugated proteins?
complex that contain non-protein component added to protein such as proteoglycan or glycoprotein
Types of proteins (3)
- Fibrous proteins
- Globular proteins
- Transmembrane proteins
What is the structural feature of globular proteins? (2)
- Polar residues face outside while hydrophobic residues face interior.
- Small cavities exist within molecule
5 detection method of peptide and protein
- specific enzyme assay
- Electrophoretic analysis
- antibodies
- dyes such as coomassie blue or bradford test
- UV absorption (aromatic rings in aa absorb UV light)
defined direction of primary structure (polypeptide)
N to C (left to right)
Different primary structure can give rise to similar folding. Give an example.
Insulin
Porcine insulin has structural similarity to human.
5 types of stabilizing forces between R groups of aa
- Electrostatic bonds between positive and negative aa
- Hydrogen bonds (Serine, Threonine etc)
- Disulfide bridge between thiol group
- Hydrophobic interactions (aromatic rings)
- Van der waals forces
secondary structure of protein
hydrogen bonding arrangement of protein backbone
Is alpha-helix structure right handed or left handed?
Right handed (clockwise)
How does hydrogen bond in alpha helix look like?
C=O of alpha carbxyl group of one aa hydrogen bonds with N-H of alpha amino group of aa 4 residues along the chain. (Intrachain)
Why is proline never used in alpha helix of secondary structure?
because it has ring structure which cause high steric hindrance.
What kind of aa not used in alpha helix? (2)
- aa that has large ring structure
- aa with strong electrostatic forces
How does hydrogen bond in beta pleated sheet look like?
carboxyl of aa hydrogen bonds with N-H of amino group of adjacent strands - interchain
What is super secondary structures?
When alpha and beta strands combine through nonrepetitive links such as Greek key, beta-meander, Beta-alpha-beta unit etc, to form a motif. Several motif can combine to form a domain where it is catalytic region.
What does non-repetitive link do in supersecondary structures?
It, made up of aa residues, links motifs and bends or turns the alpha and beta structures.
Why is proline important in non-repetitive links?
It is used to introduce bend or kinks with its large aromatic ring. When bonded to glycine, it facilitates massive kink (high steric hinderance)
What is the structure of myoglobin?
Globular protein with 8 helical region and no beta pleated sheet.
What is the purpose of quaternary structure? (3)
- increase protein stability
- for cooperativiy
- To bring catalytic sites in close proximity
How does haemoglobin depict cooperativity?
it is composed of 4 polypeptides, which interacts allosterically to increase Hb affinity for oxygen.
There are problems for protein folding such as protein dense environment in cytosol and speed of folding process. What is the solution for this?
Employ molecular chaperone such as heat shock protein
What does Hsp do and need for its job?
Assist/ensure correct folding, accelerates process
It needs ATP hydrolysis
What is the example of proteolytic processing?
Insulin
- prohormone has signal sequence which is required for delivery to ER.
Then, once in ER, disulfide bridge forms between two domain of prohormone, which leads to cleavage of c-peptide that connects two domains. Then, C-peptide and mature insulin is packaged into secretory vesicle in golgi.
How can protein deposits form?
Protein might malform for example alpha helices are replaced by beta-pleated sheets, which leads to aggregation intra-cellularly and extra-cellulary, since aggregates are proteolytic-resistant.
Where does post-translational processing occur?
at position where pairs of basic aa are located such as Arg-Arg or Arg-Lys
How does Prion disease progress?
Modifications to 2nd and 3rd structures by interacting with infectious Prion protein. In the beginning, conversion requires high activation energy which leads to slow rate. But as more and more prion molecules infect nascent non-infectious Prp molecule with exponential increase, activation energy needed for conversion is lower, leading to fast conversion. Then, infectious Prp aggregates being resistant to proteolytic cleavage.
How can Prion disease start? (2)
- Genetic as in CJD
2. Ingest of infectious Prp
How does extreme pH lead to denaturation?
lows pH disrupt bonds involving carboxyl group
high pH disrupt bonds involving amino group
What does detergent disrupt in terms of denaturation?
it disrupt hydrophobic interaction
How is disulfide bonds reduced during denaturation?
using mercaptoethanol and other reducing agent
What interaction does organic solvents denature?
hydrophobic interactions
What kinds of agent disrupts secondary structure?
Chaotropic molecule such as urea/guanidium which forms strong hydrogen bonds, thus interrupting hydrogen bond at secondary structure.
Usually denaturation is irreversible what is the exception? (2)
Pancreatic ribonuclease & thermophilic bacteria which is heat-stable.
How is protein denatured in vivo?
Through non-enzymatic modification of protein such as glycosylation or oxidation
Degree of modification can be measured by Hb as Hb is glycosylated at high level of glucose and it doesn’t affect function of Hb.
Example of protein denaturation in vivo
Oxidation of collagen which leads to aggregates
