protein evolution (homology, domains etc)

Question 1

Describe a domain?

Answer 1

Typically have their own function.

Indepently folding region.

Formed largely by hydrophobic interactions.

Tertiary structure

(important evolutionary unit)

Question 2

What percent sequence identity is the arbitrary threshold for determining if they are homologous?

Answer 2

35% sequence identity or above

Question 3

How to calculate RMSD, to determine structural similarity?

Answer 3

Align comparable proteins.

Calculate how far apart equivalent atoms are (using their c-alphas)

Use pythagoras’ theorem in 3D to do this.

Sum distances apart (square)

Divide by number of residues

(square root it)

[<3.5A is normally homologous]

Question 4

Upper threshold for determining homology from structural similarity (RMSD)?

Answer 4

<3.5A distance

Question 5

What does CATH group protein domains into?

Answer 5

Class (what major 2ndary structure present? a,ß,a/ß)

Architecture (shape of fold (barrel etc))

Topology (connectivity of fold)

**Homology **(superfamily, then family)

Question 6

What is a paralog?

What is an ortholog?

Answer 6

A paralog can be in the same species, created by a gene duplication event, and following mutations. (leading to a different but similar function, otherwise it would be same gene still)

An ortholog is in a different species, created by a speciation event! Typically same or highly similar function.

Question 7

What is the function of a Rossmann fold (motif/superfamily)?

Why important?

Answer 7

NAD binding domain. Carries out NADH dehydrogenase function, or reverse, cellular source of reducing power, but also NAD+ needs to be re-reduced itself. (NAD loses or gain Hydride H- ion)

Most common domain superfamily.

Question 8

3 multidomain proteins that have Rossmann fold, NAD binding domain?

Answer 8

Lactate and malate dehydrogenase. (transfers H- to NAD+)

And alcohol dehydrogenase (also demonstrates discontinuous dehydrogenase domain, with Rossmann fold insert in middle)

Question 9

Function of Lactate dehydrogenase?

Answer 9

Oxidises lactate to pyruvate in last step of anaerobic glycolysis, contains Rossmann fold, binding NAD+ to accept H- ion from Lactate.

C-terminal LDH catalytic domain makes it specific for oxidising lactate.

Question 10

Function of malate dehydrogenase and its evolutionary relationship to lactate dehydrogenase?

Answer 10

Oxidises malate to oxaloacetate.

Paralog of LDH (gene duplication)

Question 11

Why can sequence identity be so low in structurally similar proteins (e.g. some homologues? evolutionarily related proteins)

Answer 11

Only core and functional residues need to be conserved!

Surface residues can be altered enormously.

(Residue insertions can often occur in loops. Motif structural alignment can slightly change in response to substitutions.)

Question 12

Describe the structural consequence of single base substitution (loss of function recessive mutation) that leads to Sickle Cell anaemia:

Answer 12

Negatively charged hydrophilic Glutamate (E) changed to hydrophobic (nonpolar) Valine (V) promotes aggregation of Hb proteins.

Causing sickling crises!

Question 13

How can proteins evolve new functions?

Answer 13

Mutations (insertions, deletions, substitutions) altering folds, embellishing domains, inserting domains in other domains

Gene duplications and subsequent mutations (e.g. in globin evolution)

**Gene fusion **(connecting linked function enzymes)

Gene loss (can just lose promoter seq.) and domain rearrangement

Question 14

How can domain rearrangement occur? (creating new proteins)

Answer 14

Multi-step process, gene duplication then partial gene losses.

Question 15

How are protein kinases specific to particular reactions?

Answer 15

1) Specific amino acid sequence alters binding surface.
2) Domain architecture, same and other domains interact, (targets it to right location in cell and aid interaction)