Category Archives: Chemistry (relatively pure)

Have you had your molybdenum today?

Chemists don’t usually think of the products of a chemical reaction barreling off and penetrating another structure.   Because of the equipartition of energy, the energy of a given exothermic chemical reaction quickly gets redistributed into electronic, vibration and rotational energy and some translational energy.  It’s exactly why blasting a particular bond with exactly the right energy to break it, isn’t widely used in synthetic organic chemistry — the energy redistributes over the whole molecule too rapidly.  But that’s exactly what is thought to happen in the molybdenum storage protein according to Proc. Natl. Acad. Sci. vol. 116 pp. 26497 – 26504 ’19.

Back off a bit.  Without molybdenum we’d all be dead, as it is a critical component of the plant enzyme breaking the triple nitrogen to nitrogen (aka nitrogenase), so it can be fixed into biologic material of the plant (and ultimately us).  It takes 225 kiloCalories/mole to break N2 apart (compared to 90 kiloCalories/mole for the carbon carbon bond in ethane).

The paper concerned discusses the molybdenum storage protein of a bacterium  (Azotobacter vinelandii).  The protein is a heterohexamer of 3 alpha and 3 beta subunits with a total molecular mass of 180 kiloDaltons.

The mechanism if cleverness itself — here’s a direct quote from the abstract of the paper. “First, we show that molybdate, ATP, and Mg2+ consecutively bind into the open ATP-binding groove of the β-subunit, which thereafter becomes tightly locked by fixing the previously disordered N-terminal arm of the α-subunit over the β-ATP. Next, we propose a nucleophilic attack of molybdate onto the γ-phosphate of β-ATP, analogous to the similar reaction of the structurally related UMP kinase. The formed instable phosphoric-molybdic anhydride becomes immediately hydrolyzed and, according to the current data, the released and accelerated molybdate is pressed through the cage wall, presumably by turning aside the Metβ149 side chain. A structural comparison between MoSto and UMP kinase provides valuable insight into how an enzyme is converted into a molecular machine during evolution. The postulated direct conversion of chemical energy into kinetic energy via an activating molybdate kinase and an exothermic pyrophosphatase reaction to overcome a proteinous barrier represents a novelty in ATP-fueled biochemistry, because normally, ATP hydrolysis initiates large-scale conformational changes to drive a distant process.”

What drives the MO4 away from the ADP ? Probably electrostatic repulsion between two negative charges in the very low dielectric constant environment of the storage protein (said to be around 7 with water at 80) which does relatively little to shield the charges from each other.

Of course the SN2 reaction is like two billiard balls hitting each other with the leaving group barreling off at about the same velocity as the attacking group. How fast is that?

Pretty fast.  To figure out how fast any chemical entity is moving at 300 K (80 F) just divide 2735 by the square root of the molecular mass.  So when Iodine barrels in to methyl bromide at 243 meters second, the bromine leaves at 307 meters second.

Well the C – Br bond length  is 1.9 Angstroms, the atomic radii of Br and C are 1.8 and .7 Angstroms — So methyl bromide is 4.5 Angstroms long or 4.5 x 10^-10 meters.  So 307 meters/ second means that the bromine ion takes  roughly 10^-3 seconds to go a meter, and 10^-3 * ( 1/4.5) * 10^-10 ) seconds to go the diameter of the methyl bromide molecule.  (Of course this ignores the solvent that’s in the way impeding the Bromine anion’s progress — but that’s another story).  I put this numerology in because chemists (including me) usually don’t think about reactions this way and it’s rather humbling to do so.

How a chemical measuring stick actually works

The immune system knows something is up when a foreign peptide fragment is presented to it.  Here’s the hand holding the peptide — https://www.researchgate.net/figure/Overall-structure-of-HLA-peptide-complex_fig1_26490512.

There it sits, lying on top of a bed of beta sheets, with two side rails of alpha helices.  Proteins are big, way too big to fit into the hand, so the fragments must be chopped up into peptides no longer than 9 amino acids long (see the picture of it lying in state).

So the class assignment for today is to figure out how to design a protein which takes peptides from 10 – 16 amino acids long, and shortens them to 9 amino acids.

Obviously a trick question, because the actual amino acids making up the peptide don’t really matter much.  So somehow the protein is reacting to length rather than chemistry.

Tricky no?

ERAP1 (Endoplasmic Reticulum aminopeptidase associated with Antigen Processing has figured it out [ Proc. Natl. Acad. Sci. vol. 116 pp. 22709 – 22715 ’19 ].  It is a huge protein (948 amino acids) with four domains forming a large cavity (which it must have to accomodate a 19 amino acid paptide).  The peptide is chopped up from the amino terminal, stopping when the length reaches 9 amino acids.  The active site is at one end of the cavity, and at the other end there is a site which looks like it should cleave the carboxyterminal amino acid, but it doesn’t because the site is inactive.  However, even catalytically inactive enzymatic sites have enough structure left so they bind the substrate.

So binding of the carboxy terminal amino acid to the back site causes conformational changes transmitted through various alpha helices to the active enzyme at the other end.  It munches away removing amino acid after amino acid until the peptide gets short enough (translation 9 amino acids) so that it doesn’t push on the back site.

Incredibly clever, even though it hurts me as a chemist to see the enzyme essentially ignoring the chemistry of its substrate.

I far prefer this to politics where data is ignored.  Two examples

l. From a review of a book by Paul Krugman in the Jan/Feb 2020 Atlantic

“Krugman is substantively correct on just about every topic he addresses.” Yes except Peak Oil in 2010, Stock Market collapse in Nov 2016 and the coming recession in an article April 2019

2. Former Secretary of Labor Robert Reich in the Guardian 22 Dec ’19 — “How Trump has betrayed the working class” — by employing them and raising their wages no doubt.

How little we really understand about proteins

How little we really understand about proteins.  We ‘know’ that the 7 transmembrane alpha helices of G Protein Coupled Receptors (GPCRs) all contain hydrophobic amino acids, so they dissolve in the (hydrophobic) lipids of the membrane.  GPCRs have been intensively by chemists, molecular biologists, pharmacologists and drug chemists with the net result that as of last year “128 GPCRs are targets for drugs listed in the Food and Drug Administration Orange Book. We estimate that ∼700 approved drugs target GPCRs, implying that approximately 35% of approved drugs target GPCRs.” https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5820538/

So if you changed the hydrophobic amino acids found in the 7 transmembrane segments of GPCRs to hydrophilic ones — all hell should break loose.

Wrong says Proc. Natl. Acad. Sci. vol. 116 pp. 25668 – 25667 ’19 ].  The trick was to replace hydrophobic amino acids with hydrophilic ones with the same shape.

Thus leucine (L — single amino acid letter code) is replaced by glutamine (Q), Isoleucine (I) and Valine (V) is replaced by Threonine (T) and finally phenylalanine (F) is replaced by Tyrosine (Y).  They call this the QTY code.

Instead of destroying the structure of the GPCRs (CCR5 and CXCR4) they became water soluble, and bound their ligands CCL5 for CCR5  and CXCL12 for CXCR4 to the same extent.

Even more amazing, the QTYdesigned receptors exhibit remarkable thermostability in the presence of arginine and retained ligand-binding activity after heat treatment at 60 °C for 4 h and 24 h, and at 100 °C for 10 min.

I would never have expected this.  Would you?

Why did they even do it?  Because GPCR structures are hard to study. You either have to remove them en bloc from the membrane or dissolve them in other lipids so they don’t denature.  Why these two GPCR’s?    Because their ligands are proteins and can’t snuggle deep down inside the 7 alpha helices embedded in the membrane (they’re just too big), but bind to the outside surface.  CCL5 is an 8 kiloDalton protein (probably 80 amino acids, while CXCL12 has 93.  So just solublizing the GPCR without changing any of the amino acids external to the membrane, produces an object for study.

It would be amusing to do the same thing for a GPCR binding one of the monamines.  I doubt that they would bind, but I never would have believed this possible in the first place.

Now is the winter of our discontent

One of the problems with being over 80 is that you watch your friends get sick.  In the past month, one classmate developed ALS and another has cardiac amyloidosis complete with implantable defibrillator.  The 40 year old daughter of a friend who we watched since infancy has serious breast cancer and is undergoing surgery radiation and chemo.  While I don’t have survivor’s guilt (yet), it isn’t fun.

Reading and thinking about molecular biology has been a form of psychotherapy for me (for why, see the reprint of an old post on this point at the end).

Consider ALS (Amyotrophic Lateral Sclerosis, Lou Gehrig disease).  What needs explaining is not why my classmate got it, but why we all don’t have it.  As you know human neurons don’t replace themselves (forget the work in animals — it doesn’t apply to us).  Just think what the neurons  which die in ALS have to do.  They have to send a single axon several feet (not nanoMeters, microMeters, milliMeters — but the better part of a meter) from their cell bodies in the spinal cord to the muscle the innervate (which could be in your foot).

Supplying the end of the axon with proteins and other molecules by simple diffusion would never work.  So molecular highways (called microtubules) inside the axon are constructed, along which trucks (molecular motors such as kinesin and dynein) drag cargos of proteins, and mRNAs to make more proteins.

We know a lot about microtubules, and Cell vol. 179 pp. 909 – 922 ’19 gives incredible detail about them (even better with lots of great pictures).  Start with the basic building block — the tubulin heterodimer — about 40 Angstroms wide and 80 Angstroms high.  The repeating unit of the microtubule is 960 Angstroms long, so 12 heterodimers are lined up end to end in each repeating unit — this is the protofilament of the microtubule, and our microtubules have 13 of them, so that’s 156 heterodimers per microtubule repeat length which is 960 Angstroms or 96 nanoMeters (96 billionths of a meter).  So a microtubule (or a bunch of microtubules extending a meter has 10^7 such repeats or about 1 billion heterodimers.  But the axon of a motor neuron has a bunch of microtubules in it (between 10 and 100), so the motor neuron firing to  the muscle moving my finger has probably made billions and billions of heterodimers.  Moreover it’s been doing this for 80 plus years.

This is why, what needs explaining is not ALS, but why we don’t all have it.

Here’s the old post

The Solace of Molecular Biology

Neurology is fascinating because it deals with illnesses affecting what makes us human. Unfortunately for nearly all of my medical career in neurology ’62 – ’00 neurologic therapy was lousy and death was no stranger. In a coverage group with 4 other neurologists taking weekend call (we covered our own practices during the week) about 1/4 of the patients seen on call weekend #1 had died by on call weekend #2 five weeks later.

Most of the deaths were in the elderly with strokes, tumors, cancer etc, but not all. I also ran a muscular dystrophy clinic and one of the hardest cases I saw was an infant with Werdnig Hoffman disease — similar to what Steven Hawking has, but much, much faster — she died at 1 year. Initially, I found the suffering of such patients and their families impossible to accept or understand, particularly when they affected the young, or even young adults in the graduate student age.

As noted earlier, I started med school in ’62, a time when the genetic code was first being cracked, and with the background then that many of you have presently understanding molecular biology as it was being unravelled wasn’t difficult. Usually when you know something you tend to regard it as simple or unimpressive. Not so the cell and life. The more you know, the more impressive it becomes.

Think of the 3.2 gigaBases of DNA in each cell. At 3 or so Angstroms aromatic ring thickness — this comes out to a meter or so stretched out — but it isn’t, rather compressed so it fits into a nucleus 5 – 10 millionths of a meter in diameter. Then since DNA is a helix with one complete turn every 10 bases, the genome in each cell contains 320,000,000 twists which must be unwound to copy it into RNA. The machinery which copies it into messenger RNA (RNA polymerase II) is huge — but the fun doesn’t stop there — in the eukaryotic cell to turn on a gene at the right time something called the mediator complex must bind to another site in the DNA and the RNA polymerase — the whole mess contains over 100 proteins and has a molecular mass of over 2 megaDaltons (with our friend carbon containing only 12 Daltons). This monster must somehow find and unwind just the right stretch of DNA in the extremely cramped confines of the nucleus. That’s just transcription of DNA into RNA. Translation of the messenger RNA (mRNA) into protein involves another monster — the ribosome. Most of our mRNA must be processed lopping out irrelevant pieces before it gets out to the cytoplasm — this calls for the spliceosome — a complex of over 100 proteins plus some RNAs — a completely different molecular machine with a mass in the megaDaltons. There’s tons more that we know now, equally complex.

So what.

Gradually I came to realize that what needs explaining is not the poor child dying of Werdnig Hoffman disease but that we exist at all and for fairly prolonged periods of time and in relatively good shape (like my father who was actively engaged in the law and a mortgage operation until 6 months before his death at age100). Such is the solace of molecular biology. It ain’t much, but it’s all I’ve got (the religious have a lot more). You guys have the chemical background and the intellectual horsepower to understand molecular biology — and even perhaps to extend it.

 

The neuropharmacological brilliance of the meningococcus

The meningococcus can kill you within 12 hours after the spots appear — https://en.wikipedia.org/wiki/Waterhouse–Friderichsen_syndrome.  Who would have thought that it would be teaching us neuropharmacology.   But it is —  showing us how to make a new class of drugs, that no one has ever thought of.

One of the most important ways that the outside of a cell tells the inside what’s going on and what to do is the GPCR (acronym for G Protein Coupled Receptor).  Our 20,000 protein coding genome contains 826 of them. 108 G-protein-coupled receptors (GPCRs) are the targets of 475 Food and Drug Administration (FDA)-approved drugs (slightly over 1/3).   GPCRs are embedded in the outer membrane of the cell, with the protein going back and forth through the membrane 7 times (transmembrane segment 1 to 7 (TM1 – TM7). As the GPCR sits there usually the 7 TMs cluster together, and signaling molecules such as norepinephrine, dopamine, serotonin etc. etc. bind to the center of the cluster.   This is where the 475 drugs try to modify things.

Not so the meningococcus. It binds to the beta2 adrenergic receptor on the surface of brain endothelial cells lining cerebral blood vessels, turning on a signaling cascade which eventually promotes opening junctions of the brain endothelial cells with each other, so the bug can get into the brain.  All sorts of drugs are used to affect beta2 adrenergic receptors, in particular drugs for asthma which activate the receptor causing lung smooth muscle to relax.  All of them are small molecules which bind within the 7 TM cluster.

According to Nature Commun. vol 10 pp. 4752 –> ’19, the little hairs (pili) on the outside of the organism bind to sugars attached to the extracellular surface of the receptor, pulling on it activating the receptor.

This a completely new mechanism to alter GPCR function (which, after all,  is what our drugs are trying to do).  This means that we potentially have a whole new class of drugs, and 826 juicy targets to explore them with.

Here is one clinical experience I had with the meningococcus.  A middle aged man presented with headache, stiff neck and fever.  Normally spinal fluid is as clear as water.  This man’s was cloudy, a very bad sign as it usually means pus (lots of white blood cells).  I started the standard antibiotic (at the time)  for bacterial meningitis — because you don’t wait for the culture to come back which back then took two days.  The lab report showed no white cells, which I thought was screwy, so I went down to the lab to look for myself — there weren’t any.  The cloudiness was due to a huge number of meningococcal bacteria.  I though he was a goner, but amazingly he survived and went home. Unfortunately his immune system was quite abnormal, and the meningitis was the initial presentation of multiple myeloma.

Is the microtubule alive ??

When does inanimate matter become animate?  How about cilia — they beat and move around.  No one would call  the alpha/beta tubulin dimer from which they are formed alive.  The tubulin proteins contain 450 amino acids or so and form a globule 40 Angstroms (4 nanoMeters) in diameter.  The dimer is then 40 x 80 Angstroms and looks like an oil drum.  Then they form protofilaments stacked end to end — e.g. alpha beta alpha beta.  Then 13 protofilaments then align side by side to form the microtubule (which is 250 Angstroms in diameter, with a central hole about half that size.  Do you think you could design a protein to do this?

Lets make it a bit more complicated, and add another 10 protofilaments forming a second incomplete ring.  This is the microtubule doublet, and each cilium has 9 of them all arranged in a circle.

Hopefully you have access to the 31 October cell where the repeating unit of the microtubule doublet is shown in exquisite detail — https://www.cell.com/action/showPdf?pii=S0092-8674%2819%2931081-5. — Cell 179, 909–922 ’19

The structure is from the primitive eukaryote Chlamydomonas, the structure repeats every 960 Angstroms (e.g every 12 alpha/beta tubulin dimers).  So just for one repeating unit which is just under 1/10 of a micron (10,000 Angstroms) there are (13 + 10) * 12 = 276 dimers.  The cilium is 12 microns long so that’s 12 * 276 * 100 = 298,080 alpha tubulin dimers/microtubule doublet. The cilium has 9 of these + another doublet in the center, so thats 2,980,800 alpha tubulin dimers/cilium.

The cell article is far better than this, because it shows how the motor proteins which climb along the outside of the doublet (such as dynein) attach.The article also describes the molecular ruler (basically a 960 Angstrom coil coil which spans the repeat. They found some 38 different proteins associated with the microtubule repeat.  They repeat as well at 80, 160, 240, 480 and 960 Angstrom periodicity.  The proteins in the hole in the center of the microtubule (e.g. the lumen) are rich in a protein module called the EF hand which binds calcium, and which likely causes movement of the microtubule, at which point the damn thing (whose structure we now know) appears alive.

Because of the attachment of the partial ring (B ring) to the complete ring of protofilaments, each of the 23 protofilaments has a unique position in the doublet, and each of the proteins in the lumen is bound to a specific mitotubule profilament. There are 6 different coiled coil proteins inside the A ring, occupying  specific furrows between the protofilaments.

Staggering complexity built from a simple subunit, but then Monticello is only made of bricks.

Barking up the wrong therapeutic tree in Alzheimer’s disease

Billions have been spent by big pharma (and lost) trying to get rid of the senile plaque of Alzheimer’s disease.  The assumption has always been that the plaque is the smoking gun killing neurons.  But it’s just an assumption which a recent paper has turned on its ear [ Proc. Natl. Acad. Sci. vol. 116 23040 – 23049 ’19 ]

It involves a protein, likely to be a new face even to Alzheimer’s cognoscenti.  The protein is called SERF1A (in man) and MOAG-4 in yeast. It enhances amyloid formation, the major component of the senile plaque.  SERF1A is clearly doing something important as it has changed little from the humble single yeast cell to man.

The major component of the senile plaque is the aBeta peptide of 40 and/or 42 amino acids.  It polymerizes to form the amyloid of the plaque.  The initial step of amyloid formation is the hardest, getting a bunch of Abeta peptides into the right conformation (e.g. the nucleus) so others can latch on to it and form the amyloid fiber.   It is likely that the monomers and oligomers of Abeta are what is killing neurons, not the plaques, otherwise why would natural selection/evolution keep SERF1A around?

So, billions of dollars later, getting rid of the senile plaque turns out to be a bad idea. What we want to do is increase SERF1A activity, to sop up the monomers and oligomers. It is a 180 degree shift in our thinking. That’s the executive summary, now for the fascinating chemistry involved.

First the structure of SERF1A — that is to say its amino acid sequence.  (For the nonChemists — proteins are linear string of amino acids, just as a word is a linear string of characters — the order is quite important — just as united and untied mean two very different things). There are only 68 amino acids in SERF1A of which 14 are lysine 9 are arginine 5 Glutamic acid and 5 Aspartic acid.  That’s interesting in itself, as we have 20 different amino acids, and if they occurred randomly you’d expect about 3 -4 of each.  The mathematicians among you should enjoy figuring out just how improbable this compared to random assortment. So just four amino acids account for 33 of the 68 in SERF1A  Even more interesting is the fact that all 4 are charged at body pH — lysine and arginine are positively charged because their nitrogen picks up protons, and glutamic and aspartic acid are negatively charged  giving up the proton.

This means that positive and negative can bind to each other (something energetically quite favorable).  How many ways are there for the 10 acids to bind to the 23 bases?  Just 23 x 22 x 21 X 20 X 19 X 18 x 17 x 16 x 15 x 14 or roughly 20^10 ways.  This means that SERF1A doesn’t have a single structure, but many of them.  It is basically a disordered protein.

The paper shows exactly this, that several conformations of SERF1 are seen in solution, and that it binds to Abeta forming a ‘fuzzy complex’, in which the number of Abetas and SERF1s are not fixed — e.g. there is no fixed stoichiometry — something chemists are going to have to learn to deal with — see — https://luysii.wordpress.com/2018/12/16/bye-bye-stoichiometry/.  Also different conformations of SERF1A are present in the fuzzy complex, explaining why it has such a peculiar amino acid composition.  Clever no?  Let’s hear it for the blind watchmaker or whatever you want to call it.

The paper shows that SERF1 increases the rate at which Abeta forms the nucleus of the amyloid fiber.  It does not help the fiber grow.  This means that the fiber is good and the monomers and oligomers are bad.  Not only that but SERF1 has exactly the same effect with alpha-synuclein, the main protein of the Lewy body of Parkinsonism.

So the paper represents a huge paradigm shift in our understanding of the cause of at least 2 bad neurological diseases.   Even more importantly, the paper suggests a completely new way to attack them.

The incredible chemical intelligence of an inanimate enzyme

God, I love organic chemistry.  Here’s why.  A recent Nature paper [ vol. 573 pp. 609 – 613 ’19 ] shows that an enzyme uses a Newton’s cradle to shuttle an allosteric effect some 25 Angstroms between two catalytic centers.  I’d never heard of Newton’s cradle, but you’ll recognize it from the picture when you follow this link — https://en.wikipedia.org/wiki/Newton%27s_cradle.  It is a device used to show that most classic example of classical (e.g. nonQuantum) physics — the conservation of momentum.

This despite Feynman’s statement in the Feynman Lectures on Physics Vol I. p 12 – 6 “Molecular forces have never been satisfactorily explained on a basis of classical physics” it takes quantum mechanics to understand them fully.”  True but chemists think of reactions in terms of classic physics all the time (harmonic oscillators as bond models, billiard ball collections hitting each other as in SN2).

To understand what is going on, you must understand the low barrier hydrogen bond. [ Proc. Natl. Acad. Sci. vol. 95 pp. 12799 – 12802 ’98 ] which is a type of hydrogen bond postulated to occur in enzymes, in which the potential barrier to shifting the hydrogen from one nucleophile (oxygen or nitrogen) in the bond to another is quite low (2 Kcal/mole). The nucleophiles are closer together than they usually are ( e. g. the interatomic distance between the two heteroatoms is smaller than the sum of their van-der-Waals radii (≤ 2.55 Å for O–O pairs; ≤ 2.65 Å for O–N pairs), and the hydrogen is essentially covalently bonded to both. This makes the hydrogen bonds quite strong (10 – 20 Kcal/mole). They think that such bonds stabilize intermediates in enzymatic reactions (such as that formed by the catalytic triad of a serine protease).

Regard the low barrier hydrogen bond as what glues the balls together in the Wiki picture.

The enzyme described in the paper (transketolase) uses a chain of low barrier hydrogen bonds as a communication channel between the two remote (25 Angstroms away) active sites in the obligate functional dimers.

The still pictures have to be seen to be believed.  I can’t wait for the movie.

How general anesthesia works

People have been theorizing how general anesthesia works since there has been general anesthesia.  The first useful one was diethyl ether (by definition what lipids dissolve in).  Since the brain has the one of the highest fat contents of any organ, the mechanism was obvious to all.  Anesthetics dissolve membranes.  Even the newer anesthetics look quite lipophilic — isoflurane CF3CHCL O CF2H screams (to the chemist) find me a lipid to swim in.  One can show effects of lipids on artificial membranes but the concentrations to do so are so high they would be lethal.

Attention shifted to the GABA[A] receptor, because anesthetics are effective in potentiating responses to GABA  — all the benzodiazepines (valium, librium) which bind to it are sedating.  Further evidence that a protein is involved, is that the optical isomers of enflurane vary in anesthetic potency (but not by very much — only 60%).  Lipids (except cholesterol) just aren’t optically active.  Interestingly, alfaxolone is a steroid and a general anesthetic as well.

Well GABA[A] is an ion channel, meaning that its amino acids form alpha helices which span the membrane (and create a channel for ion flow).  It would be devilishly hard to distinguish binding to the transmembrane part from binding to the membrane near it. [ Science vol. 322 pp. 876 – 880 2008 ] Studied 4 IV anesthetics (propofol, ketamine, etomidate, barbiturate) and 4 gasses (nitrous  oxide, isoflurance, devoflurane, desflurane) and their effects on 11 ion channels — unsurprisingly all sorts of effects were found — but which ones are the relevant.

All this sort of stuff could be irrelevant, if a new paper is actually correct [ Neuron vol. 102 pp. 1053 – 1065 ’19 ].  The following general anesthetics (isoflurane, propofol, ketamine and desmedtomidine) all activate cells in the hypothalamus (before this anesthetics were thought to work by ultimately inhibiting neurons).  They authors call these cells AANs (Anesthesia Activated Neurons).

They are found in the hypothalamus and contain ADH.  Time for some anatomy.  The pituitary gland is really two glands — the adenohypophysis which secretes things like ACTH, TSH, FSH, LH etc. etc, and the neurohypophysis which secretes oxytocin and vasopressin (ADH) directly into the blood (and also into the spinal fluid where it can reach other parts of the brain.  ADH release is actually from the axons of the hypothalamic neurons.  The AANs activated by the anesthetics release ADH.

Of course the workers didn’t stop there — they stimulated the neurons optogenetically and put the animals to sleep. Inhibition of these neurons shortened the duration of general anesthesia.

Fascinating (if true).  The next question is how such chemically disparate molecules can activate the AANs.  Is there a common receptor for them, and if so what is it?

Happy fiscal new year !

A sad (but brilliant) paper about autophagy

Over the past several decades I’ve accumulated a lot of notes on autophagy (> 125,000 characters).  It’s obviously important, but in a given cell or disease (cancer, neurodegeneration) whether it helps a cell die gracefully or is an executioner is far from clear.  Ditto for whether enhancing or inhibiting it in a given situation would be helpful (or hurtful).

A major reason for the lack of clarity despite all the work that’s been done can be found in the following excellent paper [ Cell vol. 177 pp.1682 – 1699 ’19 ].  Some 41 proteins are involved in autophagy in yeast and more in man.  Many are described as ATGnn (AuTophagy Gene nn).

Autophagy is a complicated business: forming a membrane, then engulfing various things, then forming a vacuole,  then fusing with the lysosome so that the engulfees are destroyed.

The problem with previous work is that if a protein was found to be important in autophagy, it was assumed to have that function and that function only.   The paper shows that core autophagy proteins are involved in (at least) 5 other processes (endocytosis, melanocyte formation, cytokinesis, LC3 assisted phagocytosis and translocation of vesicles from the Golgi to the endoplasmic reticulum).

Experiments deleting or  increasing a given ATGnn were assumed to produce their biological effects by affecting autophagy.

The names are unimportant.  Here is a diagram of 6 autophagy proteins forming a complex producing autophagy

1 2 3

4 5 6

So 2 binds to 1, 3 and 5

But in endocytosis

1 2 3

5

form an important complex

In cytokinesis the complex formed by

2 3

5

is important.

Well you get the idea.  Knocking out 2 has cellular effects on far more than autophagy.  So a lot of work has to be re-thought and probably repeated.

Notice that all 6 functions involve movement of membranes.  So just regard the 6 proteins as gears of different diameters which can form the guts of different machines as they combine with each other (and proteins specific to the other 5 processes mentioned) to move things around in the cell.