Lecture 3: Motifs in Proteins

Question 1

How does co-operativity work for PNA?

Answer 1

The symmetry of recognition sequences means that DNA has two-half sites that can be bound by dimeric proteins.

• Inverted repeats lead to perfect two-fold symmetry.
• Having a double operator means that 30 times less protein is required than for a single operator to get to 99% bound.
• 100 times less is required to get to 99.9%.
• The nature of the half sites increases the complexity of the DNA-binding protein depending on whether they are identical or different.

Question 2

What is facilitated target location? What are the 3 ways it can occur?

Answer 2

Facilitated target location is how DNA-binding proteins seek their recognition sequences.

• Sliding. The protein slides along the DNA.
• Intersegment transfer. DNA bending can bring two sequences together.

Interdomain dissociation and association. The DNA can move off one place and bind to another

Question 3

What is the helix-turn helix motif?

Answer 3

The HTH is a common motif found in transcription regulators and enzymes.

• It is a 20 amino acid segment of two almost perpendicular α helices connected by a 4 residue β turn.
• The linker is sometimes in the form of a loop as long as the relative orientations of the α helices are maintained.
• Position 1-7 is the first helix and 12-20 is the second helix.
• The linker usually contains a glycine residue for the tight turn at position 9.
• The motif binds DNA in the major groove.
• The first helix and linker created supporting contacts with the DNA backbone.
• The second helix is the recognition helix. It is inserted into the groove and direct contacts are made between the amino acid side chains and nucleotide bases.
• Sometimes contacts are indirect and may be mediated by bridging water molecules.
• The motif is usually found in a bundle of 3 to 6 α helices that provide a stabilising core.

Question 4

What is the helix-loop-helix?

Answer 4

The HLH is a different domain to the HTH.

• They are similar to the leucine zipper.
• There is a 7-fold repeat, number 4 and 7 tend to be leucine or something similar. It is a helical coil.
• The two helices stick together like a zipper.
• b-HLH is a basic region followed by HLH.
• Sometimes you can get b-HLH-Zip.
• The basic region can interact with standard direct readout.

Question 5

How is the α-helix used in protein-nucleic acid interactions?

Answer 5

The α helix has a number of functions.

• It is well-adapted to making hydrogen bond contacts in the major groove.
• The recognition helices can widen the major groove.
• The helix is polar. The N-terminus has a ½ + charge, while the C-terminus has a ½ - charge.
• The spaces between N-Hs corresponds to the spacing between phosphates of the DNA backbone.
• The α helix can be used as a phosphate sensor.
• Backbone contacts can be used for indirect readout. They can also position the protein in a better position for creating hydrogen bonds.

Question 6

How is the β-sheet used in protein-nucleic acid interactions?

Answer 6

β-sheets create a surface of hydrogen bond donors and acceptors.

• The surfaces must have complementary hydrogen bonding and VDWs interactions.
• Antiparallel β sheets have a right-handed twist. It can form a β-antiparallel ribbon which can fit into the major groove of the DNA helix.
• The spacing between the N-Hs is fairly similar to the spacing of the phosphates. N-Hs can be used as a phosphate sensor.
• The arginine repressor from M. tuberculosis can recognise a narrow groove via phosphate contacts from a four stranded β sheet. The sheet lies above the groove without inserting side chains.