Steele - Lecture 2 Flashcards
(26 cards)
Where are disulfide bonds largely found?
- In active site of enzymes
- In extracellular environment, which is more oxidative(harsher) than the intracellular environment (reductive environment).
- -> Helps stabilize proteins in these environments
What are 2 protein motifs found in Src tyrosine-kinase?
1) glycine-rich loop: Binds phosphate of ATP
2) DFG: coordinates Mg binding to ATP
* motifs are functional/structural subunits of specific domains
What are 2 physiological examples of disulfide bonds?
1) insulin; one within alpha chain, 2 occurring between alpha and beta chains
2) antibodies: disulfide bonds link heavy to light chains and heavy to heavy chains
How are 3D structures identified?
Through X-ray diffraction pattern resulting from Fourier transformations of PROTEIN CRYSTALS
What is a method of determining protein structure for smaller proteins (<100K MW)?
NMR- apply magnetic field to proteins in solution, aligning spin states of nuclei. Energy derived from them returning to original state provides structural information.
Advantage- don’t need to crystallize.
Disadvantage- Size limit(<100K MW or 900 AA- can be used for structure of some whole viruses though)
How is quaternary structure stabilized?
multiprotein assemblies, stabilized by weak interactions like 2’ and 3’ structures
What interactions qualify as 4’ Structure?
All protein interactions COULD be considered 4’ structure, but it depends on the stability of the interaction
Can be homo or heteromeric
What is the generic formula for MW of proteins?
AA x 100 = #Da
What are two general categories of protein shape?
1) globular (dominant)
2) Structural proteins - elongated (fibrous)
What are the sources of charge for proteins?
1) amino (+) and carboxy (-) terminus)
2) side chains of basic and acidic amino acids
3) covalent modifications (eg phosphorylation) - transient
4) metal ions (eg cofactors)
How can you determine if an R group or terminus will be protonated?
Look at pKa. If,
pHpKa, deprotonated form predominates (- charge)
What is pKa?
pH at which 1/2 of ionizable group is protonated, 1/2 deprotonated
Are proteins good buffers? Why or why not?
No. A buffer is a molecule that resists pH change in solution within 1 pH unit above and below the pKa. Requires much more acid/base to change pH within buffer zone.
Proteins only have His residues and amino-terminus with pKa values within the right range (~6.5-7.5), but they are too sparse to have a significant impact.
Glycosylation and disulfide bonds are most commonly found on which side for transmembrane domains?
Extracellular domain
What modifications are found on the intracellular domain?
1) reduced sulfhydryl groups
2) phosphorylation
3) no complex carbs
What secondary structure is found in transmembrane proteins?
Strongly hydrophobic alpha helix (single-pass) or helices (multi-pass)
What are the two types of glycosylation? What AA do they modify?
1) N-linked: side chain N on asparagine
2) O-linked: OH group on serine or threonine
(don’t need to memorize)
What is Leri-Weill dyschondrosteosis caused by?
mutation in nuclear localization signal in SHOX transcription factor.
Mutations in localization signal can cause disease
What are some protein destinations in the cell?
1) ER
2) nucleus (reversible)
3) mitochondrion
4) cytoplasm (default mode)
5) peroxisome
What is a common characteristic of nuclear localization signals?
Mostly basic amino acids (arg, lys)
What is the difference between monopartite and bipartite localization signals?
Monopartite signals are continuous, bipartite signals have a sequence in the middle of the basic amino acids
(don’t need to memorize)
How are proteins isolated?
By
1) charge: ion-exchange chromatography
2) size: gel filtration chromatography
3) hydrophobicity: Absorption chromatography
4) interaction with other molecules: affinity chromatography w/ ligands, substrates, products, binding proteins, antibodies
What factors tell you if two different proteins evolved together?
Primary structure, tertiary structure (may be better index than 1’ structure), function
ex: hemoglobin, myoglobin
Define ortholog and paralog
ortholog- proteins with similar function and structure in different species
paralog- proteins with similar structure but different function in same species, often arise from gene duplication
