Lecture 17: Lummis et al.

Question 1

Where is the proline in nAchRs?

Answer 1

Between the M2 and M3 loop

Question 2

Where is serotonin a major NT?

Answer 2

Major NT in the CNS and found in the raphe nucleus

Question 3

What are some serotinergic related diseases?

Answer 3

Polymorphism in 5-HT3A gene associated with schizophrenia, depression and anxiety (SSRIs), and IBS

Question 4

Which polymorphis in the 5-HT3 receptor is associated with depression?

Answer 4

A tyr-ser polymorphism –> ppl with tyr are more prone to depression. Mutant has more prolonged, slower, activation kinetics and prolonged opening –> greater amount of channel activity, more serotonin release

Question 5

How are prolines different from normal aas in their conformation?

Answer 5

The cis and trans forms can interconvert more easily –> this may be the switch for gating

Question 6

What type of experiment did they do?

Answer 6

They did electrophysiology with site directed mutagenesis with an unnatural amino acid

Question 7

What was the main difference between the unnatural amino acids in the functional channels vs the nonfunctional channels?

Answer 7

In the functional channels, the amino acids had a large propensity to be in the cis conformation, in the non-functional channels, the amino acids had a very low propensity for cis conformation

Question 8

What did having a propensity to be in cis conformation do to gating?

Answer 8

It made gating easier, but did not change the affinity of serotonin to the binding site

Question 9

What happened to receptors with Dmp at 8*?

Answer 9

They activated with 5-HT, but did not fully deactivate. Dmp had the highest propensity to be in cis conformation (71%)

Question 10

What is the conformation of the M2-M3 peptide?

Answer 10

There are two conformations found. Major form is 5x more prevalent

Question 11

What is the major form of the receptor?

Answer 11

trans conformation of pro 8*

Question 12

What is the minor form of the receptor?

Answer 12

cis conformation of pro 8*

Question 13

What is the proposed gating mechanism of 5HT receptors?

Answer 13

Loop 2 and 7 act as calipers to lock pro 8* in the trans conformation. 5HT could bind to trpB in binding pocket, which will have an effect on the cys-loop. closing in of c loop also alters conformation. Movement of loops frees up proline to convert from trans-cis, which splays open channel

Question 14

What are the interactions with loop 2?

Answer 14

There is a salt bridge between R on b10 and E45 on M2, this interaction is important for the gating process