Watching electrons being pushed

Would any organic chemist like to watch electrons moving around in a molecule? Is the Pope Catholic? Attosecond laser pulses permit this [ Science vol. 346 pp. 336 - 339 '14 ]. An attosecond is 10^-18 seconds. The characteristic vibrational motion of atoms in chemical bonds occurs at the femtosecond scale (10^-15 seconds). An electron takes 150 attoseconds to orbit a hydrogen atom [ Nature vol. 449 p. 997 '07 ]. Of course this is macroscopic thinking at the quantum level, a particular type of doublethink indulged in by chemists all the time — http://luysii.wordpress.com/2009/12/10/doublethink-and-angular-momentum-why-chemists-must-be-adept-at-it/.

The technique involves something called pump probe spectroscopy. Here was the state of play 15 years ago — [ Science vol. 283 pp. 1467 - 1468 '99 ] Using lasers it is possible to blast in a short duration (picoseconds 10^-12 to femtoseconds 10^-15) pulse of energy (pump pulse ) at one frequency (usually ultraviolet so one type of bond can be excited) and then to measure absorption at another frequency (usually infrared) a short duration later (to measure vibrational energy). This allows you to monitor the formation and decay of reactive intermediates produced by the pump (as the time between pump and probe is varied systematically).

Time has marched on and we now have lasers capable of producing attosecond pulses of electromagnetic energy (e.g. light).

A single optical cycle of visible light of 6000 Angstrom wavelength lasts 2 femtoseconds. To see this just multiply the reciprocal of the speed of light (3 * 10^8 meters/second) by the wavelength (6 * 10^3 *10^-10). To get down to the attosecond range you must use light of a shorter wavelength (e.g. the ultraviolet or vacuum ultraviolet).

The paper didn’t play around with toy molecules like hydrogen. They blasted phenylalanine with UV light. Here’s what they said “Here, we present experimental evidence of ultrafast charge dynamics in the amino acid phenylalanine after prompt ionization induced by isolated attosecond pulses. A probe pulse then produced a doubly charged molecular fragment by ejection of a second electron, and charge migration manifested itself as a sub-4.5-fs oscillation in the yield of this fragment as a function of pump-probe delay. Numerical simulations of the temporal evolution of the electronic wave packet created by the attosecond pulse strongly support the interpretation of the experimental data in terms of charge migration resulting from ultrafast electron dynamics preceding nuclear rearrangement.”

OK, they didn’t actually see the electron dynamics but calculated it to explain their results. It’s the Born Oppenheimer approximation writ large.

You are unlikely to be able to try this at home. It’s more physics than I know, but here’s the experimental setup. ” In our experiments, we used a two-color, pump-probe technique. Charge dynamics were initiated by isolated XUV sub-300-as pulses, with photon energy in the spectral range between 15 and 35 eV and probed by 4-fs, waveform-controlled visible/near infrared (VIS/NIR, central photon energy of 1.77 eV) pulses (see supplementary materials).”

The incredible information economy of frameshifting

Her fox and dog ate our pet rat

H erf oxa ndd oga teo urp etr at

He rfo xan ddo gat eou rpe tra t

The last two lines make no sense at all, but (neglecting the spaces) they have identical letter sequences.

Here are similar sequences of nucleotides making up the genetic code as transcribed into RNA

ATG CAT TAG CCG TAA GCC GTA GGA

TGC ATT AGC CGT AAG CCG TAG GA.

GCA TTA GCC TAA GCC GTA GGA ..

Again, in our genome there are no spaces between the triplets. But all the triplets you see are meaningful in the sense that they each code for one of the twenty amino acids (except for TAA which says stop). ATG codes for methionine (the purists will note that all the T’s should be U). I’m too lazy to look the rest up, but the ribosome doesn’t care, and will happily translate all 3 sequences into the sequential amino acids of a protein.

Both sets of sequences have undergone (reading) frame shifts.

A previous post https://luysii.wordpress.com/2014/10/13/the-bach-fugue-of-the-genome/ marveled about how something too small even to be called a virus coded for a protein whose amino acids were read in two different frames.

Frameshifting is used by viruses to get more mileage out of their genomes. Why? There is only so much DNA you can pack into the protein coat (capsids) of a virus.

[ Proc. Natl. Acad. Sci. vol. 111 pp. 14675 - 14680 '14 ] Usually DNA density in cell nuclei or bacteria is 5 – 10% of volume. However, in viral capsids it is 55% of volume. The pressure inside the viral capsid can reach ten atmospheres. Ejection is therefore rapid (60,000 basepairs/second).

The AIDS virus (HIV1) relies on frame shifting of its genome to produce viable virus. The genes for two important proteins (gag and pol) have 240 nucleotides (80 amino acids) in common. Frameshifting occurs to allow the 240 nucleotides to be read by the cell’s ribosomes in two different frames (not at once). Granted that there are 61 3 nucleotide combinations to code for only 20 amino acids, so some redundancy is built in, but the 80 amino acids coded by the two frames are usually quite different.

That the gag and pol proteins function at all is miraculous.

The phenomenon is turning out to be more widespread. [ Proc. Natl. Acad. Sci. vol. 111 pp. E4342 - E4349 '14 ] KSHV (Kaposi’s Sarcoma HerpesVirus) causes (what else?) Kaposi’s sarcoma, a tumor quite rare until people with AIDS started developing it (due to their lousy immune system being unable to contend with the virus). Open reading frame 73 (ORF73) codes for a major latency associated nuclear antigen 1 (LANA1). It has 3 domains a basic amino terminal region, an acidic central repeat region (divisible into CR1, CR2 and CR3) and another basic carboxy terminal region. LANA1 is involved in maintaning KSHV episomes, regulation of viral latency, transcriptional regulation of viral and cellular genes.

LANA1 is made of multiple high and lower molecular weight isoforms — e.g. a LANA ladder band pattern seen in immunoblotting.

This work shows that LANA1 (and also Epstein Barr Nuclear antigen 1` ) undergo highly efficient +1 and -2 programmed frameshifting, to generate previously undescribed alternative reading frame proteins in their repeat regions. Programmed frameshifting to generate multiple proteins from one RNA sequence can increase coding capacity, without increasing the size of the viral capsid.

The presence of similar repeat sequences in human genes (such as huntingtin — the defective gene in Huntington’s chorea) implies that we should look for frame shifting translation in ourselves as well as in viruses. In the case of mutant huntingtin frame shifting in the abnormally expanded CAG tracts rproduces proteins containing polyAlanine or polySerineArginine tracts.

Well G, A , T and C are the 1’s and 0’s of the way genetic information is stored in our genomic computer. It really isn’t surprising that the genome can be read in alternate frames. In the old days, textual information in bytes had parity bits to make sure the 1’s and 0’s were read in the correct frame. There is nothing like that in our genome (except for the 3 stop codons).

What is truly suprising it that reading in alternate frame produces ‘meaningful’ proteins. This gets us into philosophical waters. Clearly

Erf oxa ndd oga teo urp etr at

Rfo xan ddo gat eou rpe tra t

aren’t meaningful to us. Yet gag and pol are quite meaningful (even life and death meaningful) to the AIDS virus. So meaningful in the biologic sense, means able to function in the larger context of the cell. That really is the case for linguistic meaning. You have to know a lot about the world (and speak English) for the word cat to be meaningful to you. So meaning can never be defined by the word itself. Probably the same is true for concepts as well, but I’ll leave that to the philosophers, or any who choose to comment on this.

An experiment of nature

Yesterday’s post https://luysii.wordpress.com/2014/10/15/ebola/ concerned the fact that 2 nurses taking care of a patient in Texas had been infected (presumably even after taking all the recommended precautions). Given that, I was concerned about the possibility of airborne spread.

Bryan wrote in to say the following:

“It seems doubtful airborne spread was involved. Remember, the Texas patient was initially sent home after showing symptoms, yet none of his family members were infected. Only those health workers directly involved in his care (and thus exposed to infected bodily fluids) have been infected, consistent with the idea that the disease can be transmitted only though contact with infected bodily fluids.”

I certainly hope he is right.

In something right out a novel, the possibility of airborne spread is now going to be empirically tested, as one of the two infected nurses flew to Cleveland, and then back to Texas in the 24 hours prior to her diagnosis. She apparently had a slight fever on boarding. So 100+ people were in a confined space with her for a few hours.

It’s why I don’t read fiction — reality is far more fantastic than anything writers can produce.

One more bizarre development. Here in Massachusetts, legislators today are scheduled to hear about the readiness of the state’s hospitals to handle Ebola. Amazingly, they will only get input from hospital CEOs. No nurses, thank you. Naturally the nurses are pissed as they should be (and so should you if you live in the state). If there were ever a time to hear from boots on the ground about Ebola readiness, it is now.

Addendum 17 Oct ’14

The Obama administration has just appointed a former chief of staff for former vice-president Gore and present vice-president Biden as the “Ebola czar”. Presumably, not for his medical expertise but for his ability to coordinate various governmental agencies, which was hardly the problem in the CDC’s response to the Texas cases. Hopefully, this will not be another case of “Brownie, you’re doing a heck of a job,” but I’m not optimistic — http://en.wikipedia.org/wiki/Michael_D._Brown

Now for some molecular biology. The genome of Ebola is RNA which mutates much more rapidly than DNA genomes. It does this so quickly that at death from AIDS (another RNA virus), there are so many viral variants present that the infecting ensemble is called a quasiSpecies. With a large population infected in Africa there is more Ebola virus extant than at any time in the past. There is some reason to hope that natural selection for a more transmissible form of Ebola in the large infected human population will not occur (the AIDS virus hasn’t become more infectious over the years). This is only a hope.

Ebola

This morning (15 October) it was announced that a second health care worker at the Texas hospital where an ebola patient died has ‘tested positive’ for it. If ebola can spread in a hospital environment where presumably precautions were taken, once it gets out into the populace at large it can spread much faster. This had to be human to human transmission — no other animal vector is involved (as it probably is in Africa).

How does it spread? We don’t know, but the two Texas cases probably imply that airborne spread is possible.

What to do?

In our case it means not getting into a confined space with over 100 people we don’t know from all over the world for an 8 – 16 hour period (e.g. an international flight). Have you ever been on a flight where no one had a cold?

For the USA, it should mean banning all flights from endemic countries. This has been the case in the past. My cousin’s wife has a lot of relatives in Brazil, because the people on the boat had lots of pink eye, and the boat was simply turned away over 100 years ago.

It should mean caring for Ebola patients in specialized facilities where only they are cared for –e.g. not in a general hospital since we don’t know how it spreads.

The greatest way to spread the disease (the Hajj — millions of people from all over the world crowded together for days followed by worldwide dispersal) has mercifully just ended before the disease escaped Africa to any extent.

Will ISIS or Al-Qaeda try to bring Ebola to the USA? Of course.

We live in a society where children have supervised play dates, and where walking unattended to school is almost considered child abuse. What will happen to such a risk-averse society when there is actual risk to going out to (the mall, the school, to work)?

The Bach Fugue of the Genome

There are more things in heaven and earth, Horatio,
Than are dreamt of in your philosophy.
– Hamlet (1.5.167-8), Hamlet to Horatio

Just when you thought we’d figured out what genomes could do, the virusoid of rice yellow mottle virus performs a feat of dense coding I’d have thought impossible. The following work requires a fairly sophisticated understanding of molecular biology which the articles in “Molecular Biology Survival Guide for Chemists” might provide the background. Give it a shot. This is fascinating stuff. If the following seems incomprehensible, start with –https://luysii.wordpress.com/2010/07/07/molecular-biology-survival-guide-for-chemists-i-dna-and-protein-coding-gene-structure/ and then follow the links forward.

Virusoids are single stranded circular RNAs which are dependent on a virus for replication. They are distinct from viroids because viroids need nothing else to replicate. Neither the virusoid or the viroid were thought to code for protein (until now). They are usually found inside the protein shells of plant viruses.

[ Proc. Natl. Acad. Sci. vol. 111 pp. 14542 - 14547 '14 ] Viroids and virusoids (viroid like satellite RNAs) are small (220 – 450 nucleotide) covalently closed circular RNAs. They are the smallest known replicating circular RNA pathogens. They replicate via a rolling circle mechanism to produce larger concatemers which are then processed into monomeric forms by a self-splicing hammerhead ribozyme, or by cellular enzymes.

The rice yellow mottle virus (RYMV) contains a virusoid which is a covalently closed circular RNA of a mere 220 nucleotides. A 16 kiloDalton basic protein is made from it. How can this be? Figure the average molecular mass of an amino acid at 100 Daltons, and 3 codons per amino acid. This means that 220 can code for 73 amino acids at most (e.g. for a 7 – 8 kiloDalton protein).

So far the RYMV virusoid is the only RNA of viroids and virusoids which actually codes for a protein. The virusoid sequence contains an internal ribosome entry site (IRES) of the following form UGAUGA. Intiation starts at the AUG, and since 220 isn’t an integral multiple of 3 (the size of amino acid codons), it continues replicating in another reading frame until it gets to one of the UGAs (termination codons) in UGAUGA or UGAUGA. Termination codons can be ignored (leaky codons) to obtain larger read through proteins. So this virusoid is a circular RNA with no NONcoding sequences which codes for a protein in either 2 or 3 of the 3 possible reading frames. Notice that UGAUGA contains UGA in both of the alternate reading frames ! So it is likely that the same nucleotide is being read 2 or 3 ways. Amazing ! ! !

It isn’t clear what function the virusoid protein performs for the virus when the virus has infected a cell. Perhaps there aren’t any, and the only function of the protein is to help the virusoid continue existence inside the virus.

Talk about information density. The RYMV virusoid is the Bach Fugue of the genome. Bach sometimes inverts the fugue theme, and sometimes plays it backwards (a musical palindrome if you will).

It is unfortunate that more people don’t understand the details of molecular biology so they can appreciate mechanisms of this elegance. Whether you think understanding it is an esthetic experience, is up to you. I do. To me, this resembles the esthetic experience that mathematics offers.

A while back I wrote a post, wondering if the USA was acquiring brains from the MidEast upheavals, the way we did from Europe because of WWII. Here’s the link https://luysii.wordpress.com/2014/09/28/maryam-mirzakhani/.

Clearly Canada has done just that. Here are the authors of the PNAS paper above and their affiliations. Way to go Canada !

Mounir Georges AbouHaidar
aDepartment of Cell and Systems Biology, University of Toronto, Toronto, ON, Canada M5S 3B2; and
Srividhya Venkataraman
aDepartment of Cell and Systems Biology, University of Toronto, Toronto, ON, Canada M5S 3B2; and
Ashkan Golshani
bBiology Department, Carleton University, Ottawa, ON, Canada K1S 5B6
Bolin Liu
aDepartment of Cell and Systems Biology, University of Toronto, Toronto, ON, Canada M5S 3B2; and
Tauqeer Ahmad
aDepartment of Cell and Systems Biology, University of Toronto, Toronto, ON, Canada M5S 3B2; and

The Silence is Deafening

A while back I wrote a post concerning a devastating paper which said that papers concerning the default mode of brain activity (as seen by functional magnetic resonance imaging { fMRI } ) had failed to make sure that the subjects were actually awake during the study (and most of them weren’t). The post is copied here after the ****

Here’s a paper from July ’14 [ Proc. Natl. Acad. Sci. vol. 111 pp. 10341 - 10346 '14 ] Functional brain networks are typically mapped in a time averaged sense, based on the assumption that functional connections remain stationary in the resting brain. Typically resting state fMRI (default network == rsfMRI) is sampled at a resolution of 2 seconds or slower.

However the human connectome project (HCP) has high-quality rsfMRI data at subsecond resolution (using multiband accelerated echo planar imaging. This work used a sliding window approach mapping the evolution of functional brain networks over a continuous 15 minute interval at subsecond resolution in 10 people. I wrote the lead author 21 July ’14 to ask how he knew the subjects weren’t asleep during this time.

No response. The silence is deafening.

Another more recent paper [ Proc. Natl. Acad. Sci. vol. 111 pp. 14259–14264 '14 ] had interesting things to say about brain maturation in attention deficit disorder/ hyperactivity — here’s the summary

It was proposed that individuals with attention-deficit/hyperactivity disorder (ADHD) exhibit delays in brain maturation. In the last decade, resting state functional imaging has enabled detailed investigation of neural connectivity patterns and has revealed that the human brain is functionally organized into large-scale connectivity networks. In this study, we demonstrate that the developing relationships between default mode network (DMN) and task positive networks (TPNs) exhibit significant and specific maturational lag in ADHD. Previous research has found that individuals with ADHD exhibit abnormalities in DMN–TPN relationships. Our results provide strong initial evidence that these alterations arise from delays in typical maturational patterns. Our results invite further investigation into the neurobiological mechanisms in ADHD that produce delays in development of large-scale networks.

I wrote the lead author a few days ago to ask how he knew the subjects weren’t asleep during this time.

No response. The silence is deafening.

***

If you Google “default mode network” you get 32 million hits in under a second. This is what the brain is doing when we’re sitting quietly not carrying out some task. If you don’t know how we measure it using functional mMRI skip to the #### and then come back. I’m not a fan of functional MRI (fMRI), the pictures it produces are beautiful and seductive, and unfortunately not terribly repeatable.

If [ Neuron vol. 82 pp. 695 - 705 '14 ] is true than all the work on the default network should be repeated.

Why?

Because they found that less than half of 71 subjects studied were stably awake after 5 minutes in the scanner. E.g. they were actually asleep part of the time.

How can they say this?

They used Polysomnography — which simultaneously measures tons of things — eye movements, oxygen saturation, EEG, muscle tone, respiration pulse; the gold standard for sleep studies on the patients while in the MRI scanner.

You don’t have to be a neuroscientist to know that cognition is rather different in wake and sleep.

Pathetic.

####

There are now noninvasive methods to study brain activity in man. The most prominent one is called BOLD, and is based on the fact that blood flow increases way past what is needed with increased brain activity. This was actually noted by Wilder Penfield operating on the brain for epilepsy in the 30s. When the patient had a seizure on the operating table (they could keep things under control by partially paralyzing the patient with curare) the veins in the area producing the seizure turned red. Recall that oxygenated blood is red while the deoxygenated blood in veins is darker and somewhat blue. This implied that more blood was getting to the convulsing area than it could use.

BOLD depends on slight differences in the way oxygenated hemoglobin and deoxygenated hemoglobin interact with the magnetic field used in magnetic resonance imaging (MRI). The technique has had a rather checkered history, because very small differences must be measured, and there is lots of manipulation of the raw data (never seen in papers) to be done. 10 years ago functional magnetic imaging (fMRI) was called pseudocolor phrenology.

Some sort of task or sensory stimulus is given and the parts of the brain showing increased hemoglobin + oxygen are mapped out. As a neurologist, I was naturally interested in this work. Very quickly, I smelled a rat. The authors of all the papers always seemed to confirm their initial hunch about which areas of the brain were involved in whatever they were studying. Science just isn’t like that. Look at any issue of Nature or Science and see how many results were unexpected. Results were largely unreproducible. It got so bad that an article in Science 2 August ’02 p. 749 stated that neuroimaging (e.g. functional MRI) has a reputation for producing “pretty pictures” but not replicable data. It has been characterized as pseudocolor phrenology (or words to that effect).

What was going on? The data was never actually shown, just the authors’ manipulation of it. Acquiring the data is quite tricky — the slightest head movement alters the MRI pattern. Also the difference in NMR signal between hemoglobin without oxygen and hemoglobin with oxygen is small (only 1 – 2%). Since the technique involves subtracting two data sets for the same brain region, this doubles the error.

The thermodynamic subtlety of cholera

Who knew that the cholera organism passed a thermodynamics course with flying colors? Consider that it has to function at widely different temperatures (37 C when it infects us, and 20 – 30 C when it’s out in the world). When it infects us it needs to make toxins and build a secretion system to export it. This cost a lot of metabolic money (ATP). Clearly there’s no point in doing this at temperatures outside the body and a lot of reasons not to (at least 60 as turning on toxin production and building the secretion system involves synthesizing at least 60 different proteins).

If some of the following terms are unfamiliar have a look at https://luysii.wordpress.com/2010/07/07/molecular-biology-survival-guide-for-chemists-i-dna-and-protein-coding-gene-structure/ and follow the links.

How does thermodynamics help the organism turn on these genes at body temperature (37 C in us)? ToxT is a protein which turns on production of the 60 proteins. The mRNA for ToxT is only translated into protein by the ribosome at 37 C.

[ Proc. Natl. Acad. Sci. vol. 111 pp. 14241 - 14246 '14 ] The mRNA for ToxT has what the authors call an RNA thermometer in its untranslated region. It is just a sequence of nucleotides which binds to the Shine Dalgarno (SD) element (http://en.wikipedia.org/wiki/Shine-Dalgarno_sequence) in the ToxT mRNA tying it up, so the SD element can’t bind to the ribosome, meaning the mRNA for ToxT can’t be transcribed into protein . Guess what? The thermometer only binds to the SD element at low temperatures, at higher temperatures the binding is unstable leaving the SD sequence free, turning on synthesis of ToxT which then turns on the 60 proteins involved in toxin production. Clever no?

Cholera is a terrible disease, afflicting less developed countries causing terrible infant mortality. I can’t resist mentioning a completely avoidable epidemic inflicted in the name of risk reduction years ago.

[ Nature vol. 354 p. 255 '91 ] An amazing article places the blame for the cholera epidemic sweeping South America starting in Peru on a misguided application of an Environmental Protection Study implicating water chlorination as a cause of cancer. During the 80’s Peruvian officials, citing the EPA study, stopped chlorinating many of the well in Lima. However, others say that the decision might have been more based on economics than data from the EPA.

It is comforting to know that the 3516 who have died so far have been spared a long bout with cancer.

9 Oct ’14 — Emo wrote the following comment today

Story of Peruvian officials stopping chlorinating water supply based on EPA study was debunked in a study published in Lancet one year after the nature news story: Swerdlow et al. “Waterborne Transmission of Epidemic Cholera in Trujillo, Peru: Lessons for a Continent at Risk,” Lancet Vol. 340 No. 8810 (July 4, 1992), pgs. 28-33. They never chlorinated water in Trujillo, second largest city in the country because they didn’t believe deep well water needed disinfection and cost of chlorinator and chlorine was too much

Thanks Emo

Can losing one gene do all that? Yes it can — there’s still hope

The Cancer Genome Atlas has dashed our hopes of finding ‘the’ cause of cancer. It has sequenced the genomes of a large number of cancers — the following paper looked at 21 tumor types sequencing the protein coding parts (exomes) of 4,742 specimens, along with that of normal tissues [ Nature vol. 505 pp. 495 - 501 '14 ].

The problem is that lots of mutations have been found in every type of cancer studied this way.

The following is typical — 178 cases of lung cancer (squamous cell variety) were studied. Some 360 mutations in exons, 165 genomic rearrangements, and 323 copy number alterations were found — but this doesn’t represent the results for the 178 cases as a whole. This was the average amount of genomic mayhem seen in each individual tumor . How do you find ‘the’ cause of the cancer in this mess? One way might be to find a gene mutated in all 178 cases (e. g. recurrent mutations). This would be the holy grail — the mutation driving cancer formation, the rest being the chaff of the well known genomic instability due to the high mutation rate of cancer cells. They found 11 such genes, but they were far from mutated in all cases. Pretty depressing isn’t it?

A recent paper [ Proc. Natl. Acad. Sci. vol. 111 pp. 14009 - 14010, E4066 - E4075 '14 ] gave an example of a huge number of changes in the clinical activity of a cancer cell line due to the functional loss of just one gene (called COSMC). Here’s what happened. In a pancreatic cancer cell line, COSMC knockout produced malignant xenografts (e.g. placing the cells in an immunodeficient animal and watching what happens), which could be reversed by reintroduction of COSMC. The changes include (1) increased proliferation, (2)loss of contact inhibition of growth, (3) loss of tissue architecture, (4) less basement membrane adhesion and (5) invasive growth — remarkable that knocking out just one gene could do so much. Perhaps not a driver mutation, but certainly a delicious drug target. Before getting too excited, remember that this occurred in a cell line which was cancerous to begin with.

The quick and dirty explanation of what is going on is that COSMC is a protein chaperone for an enzyme adding a sugar to proteins destined either for secretion or for insertion into the cell membrane. Lose COSMC and the whole pattern of sugar attachments to these proteins changes. There are a lot of proteins modified by adding sugars (glycosylated proteins), actually 446 of them, with 1,471 sites for this to happen.

The rest of the post is for the cognoscenti and concerns the gory details.

From the paper itself — “Neoplastic transformation of human cells is virtually always associated with aberrant glycosylation of proteins and lipids.” The most frequently seen glycophenotype are the Tn and STn carbohydrate epitopes of epithelial cell cancers. They arise when mucin-type O-linked glycans (normally more complex) are truncated so that only a single -N-acetylgalactosamine (Tn) or N-acetylgalactosamine modified with sialic acid (STn) remains attached to the protein by a serine or a threonine. There are ‘up to’ 20 GalNAc transferases adding GalNAc to serine or threonine. Overall there are some 200 glycosyltransferase found in the secretory pathway. In most cases the GalNAc is modified with beta 1 –> 3 galactose by a single enzyme (called C1GalT1). This reaction is dependent on COSMC, a protein chaperone.

Although there weren’t mutations in the glycosyltransferases studied in 46 cases of pancreatic cancer, 40% of them showed hypermethylation of the COSMC (e.g. methylated cytosines in the promoter region, which shut down transcription of COSMC). This correlated with expression of truncated O-Glycans (e.g. the Tn and STn antigens) and loss of C1GalT expression.

Maryam Mirzakhani

“The universal scientific language is broken English.” So sayeth Don Voet 50+ years ago when we were graduate students. He should know, as his parents were smart enough to get the hell out of the Netherlands before WWII. I met them and they told me that there was some minor incident there involving Germans who promptly went bananas. They decided that this wasn’t the way a friendly country behaved and got out. Just about everyone two generations back in my family was an immigrant, so I heard a lot of heavily accented (if not broken) English growing up.

Which (at last) brings us to Maryam Mirzakhani, a person probably not familiar to chemists, but a brilliant mathematician who has just won the Fields Medal (the Nobel of mathematics). Born in Teheran and educated through college there, she came to Harvard for her PhD, and has remained here ever since and is presently a full prof. at Stanford.

Why she chose to stay here isn’t clear. The USA has picked up all sorts of brains from the various European upheavals and petty hatreds (see http://luysii.wordpress.com/2013/10/27/hitlers-gifts-and-russias-gift/). Given the present and past state of the middle East, I’ve always wondered if we’d scooped up any of the talent originating there. Of course, all chemists know of E. J. Corey, a Lebanese Christian, but he was born here 86 years ago. Elias Zerhouni former director of the NIH, was born in Algeria. That’s about all I know at this level of brilliance and achievement. I’m sure there are others that I’ve missed. Hopefully more such people are already here but haven’t established themselves as yet. This is possible, given that they come from a region without world class scientific institutions. Hitler singlehandedly destroyed the great German departments of Mathematics and Physics and the USA (and England) picked up the best of them.

Given the way things are going presently, the USA may shortly acquire a lot of Muslim brains from Europe. All it will take is a few random beheadings of Europeans in their home countries by the maniacs of ISIS and their ilk. Look what Europeans did to a people who did not physically threaten them during WWII. Lest you think this sort of behavior was a purely German aberration, try Googling Quisling and Marshal Petain. God knows what they’ll do when they are actually threatened. Remember, less than 20 years ago, the Europeans did nothing as Muslims were being slaughtered by Serbs in Kosovo.

Not to ignore the awful other side of the coin, the religious cleansing of the middle East of Christians by the larger Muslim community. The politically correct here have no love of Christianity. However, the continued passivity of American Christians is surprising. Whatever happened to “Onward Christian Soldiers” which seemed to be sung by all at least once a week in the grade school I attended 60+ years ago.

These are very scary times.

Now we know why hot food tastes differently

An absolutely brilliant piece of physical chemistry explained a puzzling biologic phenomenon that organic chemistry was powerless to illuminate.

First, a fair amount of background

Ion channels are proteins present in the cell membrane of all our cells, but in neurons they are responsible for the maintenance of a membrane potential across the membrane, which has the ability change abruptly causing an nerve cell to fire an impulse. Functionally, ligand activated ion channels are pretty easy to understand. A chemical binds to them and they open and the neuron fires (or a muscle contracts — same thing). The channels don’t let everything in, just particular ions. Thus one type of channel which binds acetyl choline lets in sodium (not potassium, not calcium) which causes the cell to fire impulses. The GABA[A] receptor (the ion channel for gamma amino butyric acid) lets in chloride ions (and little else) which inhibits the neuron carrying it from firing. (This is why the benzodiazepines and barbiturates are anticonvulsants).

Since ion channels are full of amino acids, some of which have charged side chains, it’s easy to see how a change in electrical potential across the cell membrane could open or shut them.

By the way, the potential is huge although it doesn’t seem like much. It is usually given as 70 milliVolts (inside negatively charged, outside positively charged). Why is this a big deal? Because the electric field across our membranes is huge. 70 x 10^-3 volts is only 70 milliVolts. The cell membrane is quite thin — just 70 Angstroms. This is 7 nanoMeters (7 x 10^-9) meters. Divide 7 x 10^-3 volts by 7 x 10^-9 and you get a field of 10,000,000 Volts/meter.

Now for the main course. We easily sense hot and cold. This is because we have a bunch of different ion channels which open in response to different temperatures. All this without neurotransmitters binding to them, or changes in electric potential across the membrane.

People had searched for some particular sequence of amino acids common to the channels to no avail (this is the failure of organic chemistry).

In a brilliant paper, entropy was found to be the culprit. Chemists are used to considering entropy effects (primarily on reaction kinetics, but on equilibria as well). What happens is that in the open state a large number of hydrophobic amino acids are exposed to the extracellular space. To accommodate them (e.g. to solvate them), water around them must be more ordered, decreasing entropy. This, of course, is why oil and water don’t mix.

As all the chemists among us should remember, the equilibrium constant has components due to kinetic energy (e.g. heat, e.g. enthalpy) and due to entropy.

The entropy term must be multiplied by the temperature, which is where the temperature sensitivity of the equilibrium constant (in this case open channel/closed channel) comes in. Remember changes in entropy and enthalpy work in opposite directions —

delta G(ibbs free energy) = delta H (enthalpy) - T * delta S (entropy

Here’s the paper [ Cell vol. 158 pp. 977 - 979, 1148 1158 '14 ] They note that if a large number of buried hydrophobic groups become exposed to water on a conformational change in the ion channel, an increased heat capacity should be produced due to water ordering to solvate the hydrophobic side chains. This should confer a strong temperature dependence on the equilibrium constant for the reaction. Exposing just 20 hydrophobic side chains in a tetrameric channel should do the trick. The side chains don’t have to be localized in a particular area (which is why organic chemists and biochemists couldn’t find a stretch of amino acids conferring cold or heat sensitivity — it didn’t matter where the hydrophobic amino acids were, as long as there were enough of them, somewhere).

In some way this entwines enthalpy and entropy making temperature dependent activation U shaped rather than monotonic. So such a channel is in principle both hot activated and cold activated, with the position of the U along the temperature axis determining which activation mode is seen at experimentally accessible temperatures.

All very nice, but how many beautiful theories have we seen get crushed by ugly facts. If they really understood what is going on with temperature sensitivity, they should be able to change a cold activated ion channel to a heat activated one (by mutating it). If they really, really understood things, they should be able to take a run of the mill temperature INsensitive ion channel and make it temperature sensitive. Amazingly, the authors did just that.

Impressive. Read the paper.

This harks back to the days when theories of organic reaction mechanisms were tested by building molecules to test them. When you made a molecule that no one had seen before and predicted how it would react you knew you were on to something.

Follow

Get every new post delivered to your Inbox.

Join 68 other followers