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Flashcards in PIP2 Kir's Deck (22)
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1
Q

Which channels were first recognized as PIP2 dependent (who)?

A

Kir6.x (KATP) channels were the first channels recognized as PIP2 dependent (Hilgemann, 1996)

2
Q

What kind of channels are Kir6.x’s ?

A

They are members of the larger superfamily of inward rectifier channels all of which may require PIP2 for function.

3
Q

How has Kir channel PIP2 activity been shown important in humans (who)?

A

Some mutant Kir channels associated with inherited diseases have increased susceptibility to modulation by stimuli that decrease membrane PIP2 levels, exacerbating the disease phenotype (Plaster, 2001).

4
Q

How do Kir channels depend on PIP2?

A

In general, Kir channels run down if PIP2 is depleted and reactivate if PIP2 micelles are applied.

5
Q

How does PIP2 react with Kir’s?

A

PIP2 stabilizes the open state. C-terminal fusion proteins made from Kir1.1 (ROMK1), Kir2.1 (IRK1), Kir3.1 (GIRK1), or Kir3.2 (GIRK2) channels bind PIP2 directly (Huang, 1998).

6
Q

What mutagenesis study has shown us the residue underlying PIP2 affinity in Kir’s?

A

Mutating a homologous arginine in the second transmembrane segment of Kir1.1, Kir2.1, Kir5.1, and Kir6.2 reduces the apparent affinity for PIP2 (Huang, 1998).

7
Q

Is arginine the only reside implicated in PIP2 binding?

A

Other basic and hydrophobic residues in the N terminus and distal C terminus are implicated in PIP2 binding to the channels (Lopes, 2002)

8
Q

What is the significance of variations in PIP2 binding residues in Kir?

A

They may enhance the specificity and affinity toward PIP2 in Kir2.1 compared to other types of Kir channels.

9
Q

Besides amino acid composition, what influences PIP2 binding to Kir?

A

Other signals regulate apparent affinity

10
Q

What signals regulate Kir1.1 affinity for PIP2?

A

Phosphorylation of Kir1.1 channels by protein kinase A (PKA) increases the apparent affinity for PIP2, making the current less sensitive to inhibition by PIP2 antibody (Liou, 1999)

11
Q

What signals regulate Kir3.1/4 affinity for PIP2?

A

Affinity of Kir3.1/4 (GIRK1/4) channels for PIP2 is increased by the Gβγ subunits of GTP-binding proteins and by intracellular Na+ (Huang, 1998)

12
Q

What signals regulate Kir6.x affinity for PIP2?

A

PIP2 interaction with Kir6.x channels is reduced by intracellular ATP, reducing the probability of opening (Baukrowitz, 1998)

13
Q

What were the initial leads to a 3D structure of PIP2 binding sites in Kir channels?

A
  1. a crystal structure of a prokaryotic Kir channel KirBac1.1 that is inhibited by PIP2 and PIP3 (Kuo, 2003)
  2. structures of cytoplasmic domains of Kir3.1 and Kir 2.1 (Nishida and MacKinnon, 2002)
14
Q

What did initial crystal structures lead to?

A

From these, Logothetis et al. (2007) developed a homology model for the Kir3.1 channel and concluded that residues implicated in PIP2 binding and residues implicated in the actions of several other modulatory agents (Na+, Gβγ) cluster in the same region of the channel in space

15
Q

What might the region containing residues for PIP2 binding + modulatory agents confer?

A

Presumably this region closely affects the balance between open and closed channels and coordinates the actions of many gating modifiers

16
Q

What homology model of what channel was made at the same time as Logothetis’s?

A

Analogously, Haider et al. (2007) made a homology model of Kir6.2

17
Q

What did Haider manage to do with his model?

A

successfully docked inositol phosphates as surrogates for phosphoinositide head groups with the model

18
Q

What finally determined the PIP2 binding site?

A

Nishida et al. (2007) obtained the crystal structure of a chimera of the membrane portion of KirBac1.1 and the cytoplasmic portion of Kir3.1

19
Q

What did Nishida’s crystal structure show (2007)?

A

a ring-like cloud of basic residues a small distance from the putative bilayer that might bind to phosphoinositides

20
Q

Which model precisely predicts where PIP2’s may bind and where does it suggest?

A

In the homology model for Kir6.1 there are four PIP2 binding sites, each at the interface between the homotetrameric subunits where gating conformational shifts might occur

21
Q

What would PIP2 be bound to? as suggested by the Kir6.1 homology model -

A

The phosphoinositide would be bound by contact with six basic residues and some hydrogen-bond interactions (Haider, 2007).

22
Q

Where would the PI head group be and what are the implications of this on Kir?

A

The phosphoinositide head group is near its maximum distance from the hydrophobic bilayer, which might imply that it pulls like a spring on the structure (near the slide helix) and this strain assists channel opening