3-DEHYDROQUINASE (L4)

Question 1

What is the function of DHQ?

Answer 1

Part of the Shikimate pathway
Removed water from dehydroquinate and puts the 1st double bond into 6-membered carbon ring
Essential for biosynthesis of aromatic compounds

Question 2

What does type 1 DHQ do?

Answer 2

Catalyses a cis-dehydration of dehydroquinate via a covalent imine intermediate (Schiff base) only involved in biosynthesis

Question 3

What does type 2 DHQ do?

Answer 3

Catalyses a trans-dehydration via an enolate intermediate. Involved in biosynthesis or carbon and energy utlilisation

Question 4

What is the structure of Type 1 DHQ?

Answer 4

Dimers of identical subunits Mr 2700 (~250AA) or a component of multifunctional enzymes consisting of several shikimate pathway enzymes

Question 5

What is the structure of Type 2 DHQ?

Answer 5

Dodecamers of identical subunits of Mr 16000 (~150AA)

Question 6

Describe the folds of type 1 DHQ?

Answer 6

8 stranded parallel alpha/beta barrel

Question 7

Describe the folds of type 2 DHQ?

Answer 7

5 stranded parallel beta-sheet core flanked by 4 alpha-helices

Question 8

How is the substrate oriented by type 1 DHQ?

Answer 8

Lys170 in strand f is covalently linked to the product and is involved in forming the Schiff base
Negatively charged C-1 carboxylate interacts with the positively charged Arg213 (helix G) and Gln236 (loop h-H) while Glu46, Arg48 share H bond interactions with the C-4 and C-5 hydroxyl groups
Phe225 stacks parallel to substrate
Water may be essential in the reaction
Few AA bond directly with substrate

Question 9

How does T1 DHQ change when substrate is bound?

Answer 9

Arg213 moves into position to form salt bridge with substrate
Lys170 which forms the Schiff base is located on strand f

Question 10

Which residue is critical in the reaction mechanism of T1 DHQ and substrate?

Answer 10

Glu86 which interacts with His143

Question 11

Describe the dimer of T1 DHQ?

Answer 11

Helices F, G and H at dimer interface
Helices F interact symmetrically with eachother
Ala206 at start of helix G adopts strained conformation that allows neighbouring Lys207 to make salt bridge with C-term carboxylate in neighbouring dimer subunit
Arg213, key residue for carboxylate recognition, located at helix G subunit interface
Helix G contains Gly 209 and 216 provising flexibility. Arg213 is located between the 2 glycines and coordinates the substrate
Cross-linking between helices G and H stabilises dimer and holds Arg213 for substrate interaction