Ion Channels in Action

6 February 2015
Mutations Affecting Internal TEA Blockade Identify the
Probable Pore-Forming Region of a K+ Channel
SLIDE 1: (So what in the world does all this mean): In order to decipher this seemingly complex
amalgam of biological terms, we must first define what TEA stands for, and most importantly, its
function. TEA stands for tetra-ethyl-ammonium and its main function is to essentially stand at
the intracellular opening of the pore of an ion channel and block the potassium current.
Before diving into this topic of TEA and the mutations that affect the internal TEA
blockade, I should probably give you some more background about the parts of a potassium
channel to help you better understand my presentation.
-So as indicated in the title, this paper revolves around potassium channels, which are essentially
multimeric proteins. This term multimeric proteins simply means: a protein containing two or
more polypeptide chains, which we learned in the beginning of the quarter is just a chain of
various amino acids. Each of the subunits probably contributes to the lining of the pore of the
potassium channel, but it is still unclear what region of the protein actually lines the aqueous
pore. All we know so far is that several amino acid residues are known to lie in the external
mouth of the pore. So Ive just introduced a new term to you: amino acid residues, which I
should probably clarify. Amino acid residue is just a term to describe an amino acid molecule
that has lost a water molecule by becoming joined to a molecule of another amino acid. Thats
basically all there is to that.
Now that Ive given you some background and recapped info that we have learned in class so far,
I feel that it is time to dive into the meat of this paper. SO IN THIS PAPER, Yellen and
MacKinnon decided that they wanted to identify these residues that lie at the inner mouth
of the pore in order to define the area of the pore-forming region of the protein, thus
indicating the residues that may line the ion conduction pathway.
So now after this bulk of information, we have somewhat deciphered a huge chunk of the title
(Underline TEA blockade) We have defined what an Internal TEA blockade is and its
purpose(Underline Probable Pore-Forming Region) And I have also stated what Mackinnon and

Yellen hypothesize as to what clues can lead us to finding what the probable pore forming region
is, the clues being the amino acid residues at the inner mouth of the pore. And to recap, Amino
Acid Residues being merely amino acid molecules that have lost a water molecule by becoming
joined to a molecule of another amino acid. However we still have this chunk here (Underline
Mutations affecting) that I have not touched on, but will continue to touch on throughout the
discussion of this paper.
Before I continue, I sort of want to give you guys a jist of the portion of the title Mutations
Affecting Internal TEA Blockade. So according to past research, an amino acid residue that
SPECIFICALLY affects the functionality of intracellular TEA has been identified through a
process of mutagenesis; a process by which a form is changed in a stable manner, thus resulting
in a mutation. So this sort of gives you guys and insight of how the mutation process ties in with
the Internal TEA Blockade, but I will continue to clarify this process later on in the discussion.

SLIDE 2: Moving on to the meat of this paper, Mackinnon and Yellen wanted to investigate the
action of the internal TEA on the Shaker Potassium channel. (Shaker is just a type of potassium
channel that they used). So in order to study the TEA blockade, they needed to somehow
experiment without the influence of gating, which can also stop the flow of potassium. So that
being said, they used a mutant Shaker channel that does not inactivate during depolarizations.
And depolarization is just a fancy term that describes a sudden change in a cell that undergoes a
dramatic electrical charge, which in this case refers to something we have already learned known
as action potential. Heres a figure to help you understand what depolarization is:

(The indentation is hyperpolarization)

SLIDE 3: So, they prepared several mutations of the ShIR channel that introduces amino acid
changes in the SS1-SS2 region of the channel which is located between the S5 and S6 membrane
spanning sequences. DRAW THE SPANNING SEQUENCE FIGURE

So this figure that is presented here is highly significant in the understanding of hypothesis of
Makinnon and Yellen. These brilliant minds hypothesized that the SS1-SS2 region crosses the
membrane twice and in addition, included that some part of the SS1-SS2 region may extend to
the internal entryway of the pore; and the mutations within this region may affect internal TEA
blockade. This seemingly puzzling hypothesis truly ties in how mutations affecting Internal TEA
Blockade identify the probably pore forming region. And I will explain in depth how.
IN DEPTH EXPLANATION:
So this is going to be the part of the presentation where I will tie in all the concepts that I have
just spewed out, and I feel like after this tying in of ideas, you guys will have a better
understanding of this paper.
So I am now going to state again the primary purpose as stated by Mackinnon and Yellen: Yellen
and MacKinnon decided that they wanted to identify these residues that lie at the inner
mouth of the pore in order to define the area of the pore forming region of the protein, thus
indicating the residues that may line the ion conduction pathway.

So now for the breakdown, which will hopefully clarify their purpose:

According to the findings of their experiments, they have found that a specific amino acid
residue, known as amino acid residue 441, lies in the middle of the SS1-SS2 region and it
interacts with internal TEA. And to draw from the findings of the paper which Evelyn has
recently presented, the 449 position is critical in the EXTERNAL sensitivity, while 441, which I
am focusing on, is critical in the INTERNAL sensitivity. So as the amino acid residues 441 and
449 both lie within the region SS1-SS2, which interacts with TEA, it indicates that the SS1-SS2
region crosses the membrane twice and thus is associated with the ion conduction pore. To
explain further, TEAs purpose is to stand at the intracellular opening of the pore of an ion
channel and block the potassium current. So as TEA seems to critically react to the mutations
of both 449, as indicated by Evelyns papers findings, and 441, and both these amino acid
residues lie within the SS1-SS2 region, this indicates that the probable pore forming region of a
Potassium channel lies within this region as well. And to answer the final question of their posed
purpose, the section of indicating the residues that may line the ion pathway, logically there is
a short span of eight amino acids ranging from 441-449 which connects the two ends of the pore.
So this concludes my discussion of this paper as we have tied in all of the components together
of the seemingly complex title, and have identified the probable pore-forming region of a
potassium channel through the mutations that affect the Internal TEA Blockade. Thank You.