Darkthorn’s Blog

Describe the key physical features of the alpha-helix in terms of the number of backbone atoms, the number of residues in one turn and the hydrogen bond patterns.

• each hydrogen bond encloses a loop of the alpha-helix and contains 13 atoms, from a C = O group to the N – H group at the end
• one turn of the helix is made up of 10 atoms (3.6 residues)
• the hydrogen bonds that link one peptide bond to another are parallel to the axis of the helix and help stabilise the structure

Describe the key physical features of the beta-sheet in terms of the types of beta-sheets and forces that stabilise their structure. Where would each of the types of beta-sheets be likely to be found in a proteins structure and why?

• beta-sheets are made of multiple polypeptide strands
• hydrogen bonds form between each chain of the beta-sheet
• typically parallel beta-sheets have at least five strands, each running in the same direction. Hydrophobic residues are present only on one side. Therefore, parallel beta-sheets would be found in the middle of the protein as both sides would avoid interacting with water.
• anti-parallel beta-sheets can have as few as two strands, running in opposite directions. Hydrophobic residues are found only on one side. Therefore, anti-parallel beta-sheets would be found towards the outside of the protein, with the hydrophobic residues orientated towards the inside of the protein.

Describe the atoms in a peptide chain that form a beta-turn (specify types of atoms and relative location in the chain) and the amino acid residues commonly found in beta-turns. For each, explain why they are found in beta-turns. Also name at least one important role for beta-turns in protein structure and tertiary structure.

• a beta-turn is a tight loop formed when the carbonyl oxygen of one residue forms a hydrogen bond with the amide group of an amino acid three residues down the chain
• Proline is found in beta-turns because its cyclic structure is well suited.
• Glycine is found in beta-turns because it is the most sterically flexible (it has the smallest side chain of all amino acids)
• Beta-turns allow the formation of anti-parallel beta-sheets, which typically form towards the outside of a protein

Describe the structural features of Haemoglobin which explain its function as a cooperative binding protein.

• has two alpha subunits and two beta subunits
  • each subunit has 8 helical segments
• a cleft is formed between the E and F helices (as well as hydrophobic residues) which holds the heme group
• The negative propionate groups on the heme are stabilised by the arginine residues
• Phe 97 is packed tightly next to the heme (but not bound) when iron is situated well away from the heme group. When the ligand binds to iron, the iron is brought to interact with the heme (as well as His 101).
• The histosine removes the Phe 97 from the heme pocket into the dimer interface. The heme then settles further into the subunit.
• The conformations of two F helix residues (Lys 96 and Asp 100) are influenced by the heme being located further into the sub unit. These then link the heme with a helix on a nearby subunit, influencing the binding of a second subunit.

Describe the structure and function of the two domains of equine live alcohol dehydrogenase (ADH). Include in your answer the location and relative orientation of the substrate, the cofactor NAD and the zinc atoms. What is the effect of the NAD(H) analogue that is bound in this structure and how does it exert its effect, in terms of structural alterations at the active site?

• NAD+ domain
  • Each monomer contains a six-stranded parallel beta-sheet flanked by alpha-helixes on both sides
• Catalytic domain
  • Consists of a network of anti-parallel beta-sheets and four helixes
  • Determines the events of catalysts and the specificy of the substrate
• these two domains are separated by a deep cleft in which sit the substrate and nicotinomide moicty of NAD+
• the substrate is bound into a pocket lined with hydrophobic residues
• Structural zinc ion
  • located distant from the active site
  • involved in the folding of the enzyme
  • coordinated by four cys residues
• Catalytic zinc ion
  • located at the bottom of the active site pocket
  • coordinated by two cys residues, a histodine and the oxygen of the alcohol substrate
• CNAD (analogue) bids in the cofactor binding cleft (an additional pocket created by hydrophobic residues
  • binds to the catalytic zinc, preventing coordination to substrate
  • substrate is displaced and Ser 48 points away from the catalytic zinc, preventing hydride transfer

Describe the secondary structure of glycogen phosphorylase (GP) and the type of reaction that it catalyses. What are the features of the active site? What happens to the protein conformation upon the binding of the substrate (glucose) or an inhibitor.

• four subunits
  • 38 alpha-helices
  • 7 beta-sheets
• catalyses the phosphoric cleavage of glycogen between its alpha-1, 4-linked glucose units to produce alpha-D-1-phosphate
• binding site is formed by residues and PLP
• phosphate ion binds between the tetrazole ring of NJT and the S’-phosphate of PLP.
• When NJT binds, Leu 136 shifts to accommodate the tetrazole right and His 377 shifts to stablise the transition state of the NJT
• The loop blocking the catalytic site shifts, replacing the acidic Asp 284 residue with the basic residue Arg 569
• The Arg 569 and also Lys 568 and Lsy 574, create a high affinity phosphate substrate recognition site and electrostatic pocket for the PLP S’ phosphate group
• Aromatic compounds can bind between the aromatic rings of Phe 285 and Try 613 blocking access to the binding site.