Chpt 10 ATCase

Question 1

Aspartate Transcarbamoylase (ATCase)

Answer 1

1) Example of Allosteric Regulation
- commited step in multistep pathway to synthesize pyrimidines

2) Multisubunit enzyme 2C3 + 3R2-> C6R6
- 2 catalytic subunits; C3=trimer
- 3 regulatory subunits; r2=dimer

3) Active Site between subunits of catalytic trimers
- one subunit vs adjacent subunit

4) Exists in Two state
- alternates between T and R state

5) Bonding of substrate causes conversion from T to R state
6) Binding of CTP stabilize the T state
7) exhibits concerted model of cooperatively

Question 2

ATCase: Example of allosteric regulation

Answer 2

Committed Step in multi step pathway to synthesize pyrimidines

Question 3

ATCase: Multisubunit enzyme

Answer 3

2C3+ 3R2-> C6R6

2 Catalytic subunits:

• C3=Trimer
  a) 2 c subunits stack on top of one another
  b) each subunit contains 3 active sites
• Purified C subunit exhibits catalytic activity but unresponsive to CTP
  a) Obeys Michaelis Menten Kinetics (hyperbolic Curve)

3 regulatory subunits:

• R2=dimer
  a) binds 2 CTPs (allosteric Inhibitors); stablizes T form
  b) contains 2 Zn2+ binding sites

Each subunit contacts 2 R subunits at a structural domain in the R subunit that is stabilized by Zn2+ ionic bond to 4 cysteine residues

Question 4

ATCase: Active site

Answer 4

Active site between subunits of catalytic trimer

• one subunit: 2 Thr and Arg; 1 His and Gln
• adjacent subunit: ser and Lys

Question 5

ATCase: Exists in two states

Answer 5

Exists in two states: Alternates between T and R states
T state:
-low affinity for substrate
-CTP inhibits transition to R state

R state:
-higher affinity for substrate (cooperativity)

At equilibrium without substrate or CTP
-200 T state: 1 R state

Question 6

ATCase: Binding of Substrate

Answer 6

Binding of substrate causes conversion from T to R state
Substrate:
-Carbamoyl phosphate/aspartate or PALA
-C subunits move 12 A apart and rotate 10 degrees
-R subunits rotate 15 degrees

Question 7

ATCase: Binding of CTP

Answer 7

Binding of CTP to domain of R subunit that does not interact with C subunit stabilizes the T state (low affinity of substrate)
-Shifts E toward T state; thus inhibiting E activity