Tag Archives: MacKinnon

How one membrane protein senses mechanical stress

Chemists (particularly organic chemists) think they’re pretty smart. So see if you can figure out how a membrane embedded ion channel opens due to mechanical stress. The answer is to be found in last week’s Nature (vol. 516 pp. 126 – 130 4 Dec ’14).

As you probably know, membrane embedded proteins get stuck there because they contain multiple alpha helices with mostly hydrophobic amino acids allowing them to snuggle up to the hydrocarbon tails of the lipids making up the lipid bilayer of the biological membrane.

The channel in question is called TRAAK, known to open in response to membrane tension. It conducts potassium ions. The voltage sensitive potassium channels have 24 transmembrane alpha helices, 6 in each of the tetramer proteins comprising it. TRAAK has only 8. As is typical of all ion channels, the helices act like staves on a barrel, shifting slightly to open the pore.

In this case, with little membrane tension, the helices separate slightly permitting a a 10 carbon tail ( CH3 – [ CH2 – CH2 – CH2 ]3 – ) to enter the barrel occluding the pore. Tension on the membrane tends decrease the packing of hydrocarbon tails of the membrane, pulling the plug out of the pore. Neat !! ! ! This is a completely different mechanism than the voltage sensing helix in the 24 transmembrane voltage sensitive potassium channels, and one that no one has predicted despite all their intelligence.

Trigger warning. This paper is by MacKinnon who won the Nobel for his work on potassium channels. He used antibodies to stabilize ion channels so they could be studied by crystallography. Take them out of the membrane and they denature. Why the warning? In his Nobel work he postulated an alpha helical hairpin paddle extending outward from the channel core into the membrane’s lipid interior. It was both hydrophobic and charged, and could move in response to transmembrane voltage changes.

This received vigorous criticism from others, who felt it was an artifact produced by the use of the antibody to stabilize the protein for crystallography.

Why the warning? Because MacKinnnon also used an antibody to stabilize TRAAK.

The whole idea of membrane tension brings up the question of just how strong van der Waals forces really are. Biochemists and molecular biologists tend to think of hydrophobic forces as primarily entropic, pushing hydrophobic parts of a protein together so water would have to exquisitely structure itself to solvate them (e.g. lowering the entropy greatly). Here however, the ‘pull’ if you wish, is due to the mutual attraction of the hydrophobic lipid side chains to each other, which I would imagine is pretty week.

I’m sure that these forces have been measured, and years ago I enjoyed reading about Langmuir’s work putting what was basically soap on a substrate, and forming a two dimensional gas which actually followed something resembling P * Area = n * R * T. So the van der Waals forces have been measured, I just don’t know what they are. Does anyone out there?

Nonetheless, some very slick (physical and organic) chemistry.

Is concentration meaningful in a nanoDomain? A Nobel is no guarantee against chemical idiocy

The chemist can be excused for not knowing what a nanodomain is. They are beloved by neuroscientists, and defined as the part of the neuron directly under an ion channel in the neuronal membrane. Ion flows in and out of ion channels are crucial to the workings of the nervous system. Tetrodotoxin, which blocks one of them, is 100 times more poisonous than cyanide. 25 milliGrams (roughly 1/3 of a baby aspirin) will kill you.

Nanodomains are quite small, and Proc. Natl. Acad. Sci. vol. 110 pp. 15794 – 15799 ’13 defines them as hemispheres having a radius of 10 nanoMeters from channel (a nanoMeter is 10^-9 meter — I want to get everyone on board for what follows, I’m not trying to insult your intelligence). The paper talks about measuring concentrations of calcium ions in such a nanodomain. Previous work by a Nobelist (Neher) came up with 100 microMolar elevations of calcium in nanodomains when one of the channels was opened. Yes, evolution has produced ion channels permeable to calcium and not much else, sodium and not much else, potassium and not much else. For details read the papers of Roderick MacKinnon (another Nobelist). The mechanisms behind this selectivity are incredibly elegant — and I can tell you that no one figured out just what they were until we had the actual structures of channels in hand. As chemists you’re sure to get a kick out of them.

The neuroscientist (including Neher the Nobelist) cannot be excused for not understanding the concept of concentration and its limits.

So at a concentration of 100 microMolar (10^-4 molar) how many calcium ions does a nanoDomain contain? Well a liter has 1000 milliliters and each milliliter is 1 cubic centimeter (cc.). So each cubic centimeter is 10^7 nanoMeters on a side, giving it a volume of 10^21 cubic nanoMeters. How many cubic nanoMeters are in a hemisphere of radius 10 nanoMeters — it’s 1/2 * 4/3 * pi * 10^3 = 2095. So there are (roughly) 5 * 10^17 such hemispheres in each cc.

How many ions are in a cc. of a 1 molar solution of calcium — 6 * 10^21 (Avogadro’s #/1000). How many in a 10^-4 molar solution (100 microMolar) — 6 * 10^17. How many calcium ions in a nanoDomain at this concentration? Just (6 * 10^17)/(5 * 10^17) e.g. just over one ion/nanodomain.

Does any chemist out there think that speaking of a 100 microMolar concentration in a volume this small is meaningful? I’d love to be shown how my calculation is wrong, if anyone would care to post a comment.

They do talk about nanodomains of radius 30 nanoMeters, which still would result in under 10 calcium ions/nanoDomain.

Addendum 10 Oct ’13

My face is red ! ! ! “6 * 10^21 (Avogadro’s #/1000)” should be 6 * 10^20 (Avogadro’s #/1000), making everything worse. Here’s how the paragraph should read.

How many ions are in a cc. of a 1 molar solution of calcium — 6 * 10^20 (Avogadro’s #/1000). How many in a 10^-4 molar solution (100 microMolar) — 6 * 10^16. How many calcium ions in a nanoDomain at this concentration? Just (6 * 10^16)/(5 * 10^17) e.g. just over .1 ion/nanodomain. As Bishop Berkeley would say this is the ghost a departed ion.

Even if we increased the size of the nanoDomain by an order of magnitude (making it a hemisphere of 100 nanoMeters radius), this would give us just over 10 ions/nanodomain.