The Half Decent Pharmaceutical Chemistry Blog

Frankly, this is not the “scientific” post I thought I would have written before going on holiday, but real life has drawn my attention to a different subject: headphones. Although I haven’t begun packing yet, today I selected the headphones I will use while cycling (the main activity I’ll practice on this approaching holiday). Obviously I didn’t go for those Apple give you with the player as they are quite possibly the worst headphones I’ve ever seen.

To be frank with you, none of those I own, though, completely satisfies me: none of them, in fact, can be literally plugged in my ears. Actually I went to a shop to see how much in-ear headphones eventually cost. As it turned out, too much, at least considering the good ones (those that last more than, say, a year).

All this made me think of the role headphones play in our life. Vital, for what concerns me, as I’m not sure I could ride a bicycle without listening to music any more.
However, if you’re thinking that I’m the only headphones-iPod addict (or iPodaholic, still not sure which is the best expression), you’d probably ignore what Rad50 is. And if you do it, this means you know little about structural biology, as this protein is likely to be the best-known member of the outrageously famous ABC-ATPase superfamily.

This protein is, indeed, dramatically important: it repairs DNA double-strand breaks once it has dimerized, an event that occurs only in the presence of ATP. That’s due to a particular, striking feature of Rad50. Each monomer is formed by two lobes: one contains the Walker A sequence, while the latter the Walker B and the Signature Motif. ATP, however, binds to a site created by the Walker A and B of a monomer, whereas the Signature Motif comes from other one: this not only strengthens the interaction with ATP (as the distances are excellent for this purpose), but it also stabilizes the dimmer.
At this point you might be wondering what connection between headphones and such a protein one could possibly find. Well, the answer is simple: although structural biologists do not entirely agree on this matter, the most plausible dimerization model to date is the so-called single dimmer headphone one. According to this theory, the two globular heads of the dimmer would be very close to each other, when bound to ATP.
However, given that the protein dimerizes only in the presence of ATP, when this molecule is absent the two monomers really look like headphones: two globular heads, linked by a wire which, in this case, is a coiled coil. Each Rad50 monomer, in fact, presents an approximately 800 Å long tail: the two coils tightly wind around each other, strengthening the interaction between the monomers so that, even when ATP is missing, they are not really separated and can quickly rejoin to repair a damaged DNA.

What’s more a coiled coil is exactly how your earphones look like when you take your iPod out of your pocket, even if you had carefully and neatly wrapped them around the player. How frustrating is it?!

Since I published an email address (if you hasn't noticed it, it's right below the recent posts), some people have sent me mails, after April 3, about the French train which smashed the speed record on conventional rails.

In their letters, people asked me if the TGV V150 is faster than the TGE sequence of the Sarco/Endoplasmic Reticulum Ca-ATPase (SERCA1a).

Now, this is obviously the most pointless of all the questions, since the TGV ran on rails among stations in France, while TGE hydrolyzes a mixed anhydride, which results from the transphosphorilation of a molecule of ATP on a molecule of aspartic acid (D351).

Nevertheless, I can't let my readers down so, there we are: here comes the V150, pride of French engineering.

This has two engines, which develop a mind-blowing 25,000 horsepower. As a result this monster does 356 mph! Much more than any F1 car or Bugatti and pretty much exactly the same of a "World War II Spitfire fighter at top speed".

On the other hand, there's a protein. Welcome everyone to this spectacular P-class ATPase: it uses the hydrolysis of ATP to carry two calcium ions from the cytoplasm to the sarcoplasmic reticulum.
Actually, it doesn't hydrolyze ATP but phosphorilates an amino acid (D351), which yields a very negative deltaG. So, to sum up, it's still a proper primary active transport.

In a not-so-recent article, turnover rates of the different types of SERCAs have been determined, as well as those of the different steps.



I'm not interested in sorting out if SERCA is faster than TGV, because this would be even more meaningless (there's not even any similarity in the names), but I want to focus on the said motif.

The picture below shows how the protein changes during a complete catalytic cycle.



Briefly, the phosphorilated protein has the perfect structure to release the ions in the SR. Then, the anhydride is hydrolyzed and the whole cycle starts again.

The TGE is on the A-domain which, ehm..., just rotates on its axis, opening the pore for the ions and brining the three amino acids close to the phosphate.

Predictably, the glutamate is the most important group: using fluoroaluminate and related compounds to simulate the transition state, it has been proved that the acid activates a molecule of water (present in the crystals too) which removes the phosphate.



So, from the beginning of the cycle, the A-domain (and so the TGE motif) rotates anticlockwise of 30°, 110° on the minor axis of this cylindrical subunit, 30° up (towards the anhydride) and, finally, 110° on its minor axis again, to reach its start-position.

As you can see, the turnover rate for the dephosphorilation is 0.003 per second.

So, can we compare a train to a group of amino acids? Well, why not?

Maybe, this is not the newest theme but it's for me. This week, in fact, I read, for the very first time in my life, an article written by a Nobel Prize winner. Actually, this article is likely to be the one, that which conviced the judges.

Moreover, it deals with something that has never been covered here but, apparently, has much more to do with chemistry than I've ever thought: crystallography.

Although controversial, Roderick MacKinnon's hypothesis on "voltage-sensor paddles" is really cool.

Kayak-freaks like me suddenly get excited when hear of such a thing: voltage-dependent (potassium) channels are controlled by this hydrophobic helix-turn-helix structure, which, undoubtedly, looks like a paddle.



Controversy is something I really like: the conventional theory describes voltage sensors as a helix moving up and down inside the protein. Pretty much like a piston: not that hard to imagine, is it?
The paddle, on the contrary, can move freely through the membrane, as, mmm, a paddle in the water...



When the paddle is up, this stretches the helices that keep the pore closed. On the contrary, when the channel is closed, the sensor is there, surrounded by the phospholipids.
To sum up, this structure has the same behaviour of hydrophobic ions.

Actually, it's not that hard to believe in such a thing: valinomycin, for example, carries ions through the membranes too.

Now at this point you might be thinking: "Yes, it's all very interesting, but what about crystallography?"



Pharmaceutical chemists have an enormous problem: the lack of crystallographic pictures of receptors. That's caused by the great amount of detergent (used to remove the protein from the cell) which, predictably, remains around the transmembrane structure.

This very problem was experienced whenever people wanted to get a crystal of a voltage gated channel as well. Anyhow, it's possible to get decent results, literally, increasing the size of what's outside the membrane.
This could be achieved with the help of antibodies, binding to an epitope on the protein.

To sum up, a bright example of how crystallography can actually improve our life...and give non-chemists the opportunity to win the Noble Prize in chemistry.

Well, half prize...but this is a half-decent blog, so, it's even better, isn't it?