Wednesday, September 25, 2013

RNA Takes First Place

Biology concepts – nucleic acids, DNA, RNA, central dogma of molecular biology, ribozyme, RNA world hypothesis

The Library of Congress in Washington DC was designed as a 
showplace as well as a repository. The main reading room looks as
much like a museum or a cathedral as it does a library. If I could
figure out how to get away with it, I would live in the LOC.
Did you know that there are more than 155.3 million informational items (books and such) in the Library of Congress? Established in 1800 with 3000 volumes, the library was originally housed in the Capitol Building. Unfortunately, all the books were lost when the British fired Washington in 1814. No worries, the LOC then purchased Thomas Jefferson’s personal library of over 6500 books and set up shop in new building, although not the 1892 designed library that exists today (left).

In a way, you can think of the molecular workings of the cell like the Library of Congress. You need information storage – these are the books. In each book (chromosome or parts of a chromosome) contain the instructions (genes) needed to make products (proteins) the cell may need.

Each time you want to make a certain molecule, you must consult the book (chromosome) that has the correct instruction page (DNA gene). But you may be making many copies of your product in a short period, so one book might not be enough.

You could keep many copies of each book, maybe thousands, but this would take up too much room. The LOC already covers 2.1 million sq. feet (and that’s just one main building). What if you needed 1500 copies of One Good Turn (and interesting book about the history of the screw and screwdriver) because at some time or another, 1500 people wanted to learn how to build a square screwdriver?

To avoid this need for extra space, you make copies of pages (mRNA) from the books (chromosomes) that can be taken out of the library (nucleus) and used for making the products. Each time you want a product, a translator (tRNA and ribosome) must be used. This converts the copied instructions (mRNA) into a usable product (protein).

When one or several translations have been made, the copied instructions start to tear and get worn, and finally break down. Good thing we still have the original copy of the book stored in the nucleus… I mean library. We can go back and make more copies later if we need them. Humans are amateurs, we only have about 25,000 sets of instructions stored in 46 books, nowhere near the 155.3 million of the LOC.

The central dogma of molecular biology says that DNA is replicated to
DNA, so daughter cells get a full set of instructions. DNA is also
transcribed to mRNA, which is a copied message of the instructions to
build one protein. Finally, the mRNA acts as a code that is translated
into an amino acid polymer – a protein. HIV and other retroviruses
laugh at the central dogma, going the opposite direction, RNA to
DNA. Retrotransposons laugh at HIV, as they can do all that and more.
Cells take this library/nucleic acid analogy further. Sure, they have DNA, mRNA, and tRNA so that they can carryout the central dogma of molecular biology --- DNA goes to mRNA goes to protein (via tRNA and rRNA), but they have so much more. Just as there are many kinds of information storage at the LOC--- books, images, recordings, manuscripts, pamphlets, there are different kinds of nucleic acids as well.

Ever here of small nuclear RNAs, or micro RNAs, or plasmid DNAs for that matter? We have talked about plasmids as extrachromosomal pieces of DNA that can code for genes, especially antibiotic resistance genes in prokaryotes.

But the list of RNAs is far more impressive. There are regulatory RNAs that control gene expression (whether or not a protein is made from a gene), RNAs that control modification of other RNAs or work in DNA replication. There are even RNAs that are parasitic, like some viral genomes (RNA viruses) and retrotransposons.

Of these, retrotransposons may be the most interesting. A transposon is a piece of DNA that can jump around from place to place in the chromosomes of a cell. Barbara McClintock won a Nobel Prize for identifying transposable elements were responsible for the different colors of corn kernels in maize.

Ancient viral RNA got inserted into plant and animal genomes. The
retrotransposon can be transcribed to mRNA, and then could be
reverse transcribed back into DNA or translated into protein. The
DNA can then insert itself anywhere in the genome. Since several
mRNA transcripts can be made from one transcribed retrotransposon,
and since several pieces of DNA can be reverse transcribed from just
one mRNA, we have the potential for millions of retrotransposons in
the genome – and that’s exactly what we have found. The bottom
cartoon shows HIV. Since reverse transcription makes more mistakes
than DNA replication, many more mutants can be produced. This is
one reason HIV is so hard to treat – it’s always changing.
Retrotransposons use the library analogy to fill the shelves with hundreds of copies of themselves. If plant nuclei were like libraries, up to 80% of their book pages would be retrotransposons!

In and of themselves, retrotransposons represent an exception in nucleic acids. They are mRNA sequences that can turn back into DNA. Transcription is the process of using DNA to produce an mRNA, so going the opposite direction is called reverse transcription. This is also what retroviruses like HIV do.

In the case of retrotransposons, the chromosome held copies will be transcribed to an mRNA, and some of those copies might be translated into protein. Other copies will be reverse transcribed back to DNA by an enzyme called reverse transcriptase and will insert themselves somewhere in the genome (see picture).

In this way, retrotransposons can make more copies of themselves and end up all over the chromosomes of the organism. Mutation occurs at a higher rate in reverse transcription than in DNA replication because reverse transcriptase makes more mistakes than replication enzymes. This is why HIV is so hard to treat; it mutates so often that drug design can’t keep up with the changes in the viral proteins.

So how can the same mRNA sometimes be translated, and other times end up in a new place on the DNA? A 2013 study has investigated how one type of retrotransposon manages these different outcomes. The BARE retrotransposon of plants has just one coding sequence for a protein, but the study results show that it actually makes three distinct mRNAs from this one piece of DNA.

Sam Kean is the author of The Violinist’s Thumb, a very readable
book on molecular biology. He goes through how fruit flies were
recruited to disprove DNA heredity and ended up as the strongest
evidence for it; how DNA is linked very strongly to linguistics and
math; and how Stalin tried to breed a race of half human - half
chimps. This is in addition to showing how most DNA on Earth is
descended from viruses.
One transcript (mRNA) is modified so it can be translated but cannot be reverse transcribed. The second transcript is packaged in small bundles to be reverse transcribed later back to DNA. The third transcript type is smaller and actually houses the bundles of mRNAs to be reverse transcribed. So this retrotransposon balances itself between making protein and inserting itself into new places in the genome.

If plants have so much nucleic acid in the form of retrotransposons, could these be the remnants of ancient viral infections? You betcha, and it doesn’t stop with plants. In his fascinating book, The Violinist’s Thumb, Sam Kean lays out a compelling argument that most human DNA is actually just viral nucleic acid remnants, much of it being mutated versions of old RNAs.

Old RNA is probably the best way to describe all nucleic acids, because the generally accepted view of the evolution of life on Earth is that everything started with RNA. This called the RNA world hypothesis and professes that the job that DNA does now was first done by RNA.

The hypothesis also says that what those that protein enzymes now do - cutting things up, putting things together, and modifying existing structures - was originally done by RNAs as well, called catalytic RNAs.

We have evidence for this hypothesis, specifically, we know of many RNAs that have enzymatic activity. Called ribozymes (a cross between ribo for RNA, and zyme for enzyme), some RNAs carry out enzymatic roles in our cells and the cells of every eukaryote and prokaryote ever analyzed for their presence.

Ribozymes, a form of catalytic RNA, are present in most cells. They come
in two flavors based on what someone thought their secondary structure
looked like – the hammerhead or the hairpin. Scientists aren’t the most
imaginative when it comes to naming things. They both sit down on an
RNA where they recognize their specific sequence, and make a cut in the
strand. In the cartoon, N stands for any nucleotide, and X stands for
unknown. On the right side is a diagram showing how one ribozyme can
act again and again to cleave RNAs.
So now we are aware of two exceptions when it comes to the central dogma of molecular biology and RNA – 1) RNA can be converted back into DNA and 2) RNA can act like an protein enzyme.

One essential ribozyme function is the synthesis of protein. The ribosome (a riboprotein because it is made up of many RNAs and proteins) translates the codons of mRNA into a sequence of amino acids. It uses the RNA to link the individual amino acids together via peptide bonds. I’d say that’s essential.

Other ribozymes work on themselves. Many mRNAs, when first copied from DNA have sequence within them that is not used in the final product. These are called intervening sequences (or introns), and are cut out (spliced) as part of the transcript processing. Group I and II introns are self-splicing. They fold over on themselves and cause their own excision from the RNA of which they are part!

Group I introns can be found in the mRNAs, rRNAs, and tRNAs of most prokaryotes and lower eukaryotes, but the only place we have found them so far in higher eukaroytes are the introns of plants and the introns of mitochondrial and chloroplasts genomes.  Yet more evidence for the plastid endosymbiosis hypothesis.

If the RNA world hypothesis is to be strengthened, we must find a catalytic RNA that can replicate long strings of RNA “genes.” If RNA was both the storage material and the enzymatic material, there must have been an RNA-dependent, RNA polymerase that was itself a piece of RNA. An RNA replicase has not been found, probably because life moved on to using DNA as the long-term repository of genetic information, But we should be able to make an RNA replicase as a proof of concept.

The RNA world hypothesis is an idea of how early life on Earth transmitted
information and carried out functions. RNA did everything, stored info.,
replicated itself, and carried out enzymatic activity. A – E represent a
possible sequence, although no times can be assigned yet. According to this
theory – the last thing that developed was enzymatic proteins – but new
evidence suggests that proteins were important for the development of
tRNAs so they must have been around earlier. Step B is an area of interest,
as scientists are trying to make an RNA that could replicate any RNA, even itself.
A few ribozymes can polymerize a few nucleotides into short RNAs. The problem is that we need to show that there is an RNA that could replicate long strings of RNA that could then go on to have biological function. Until 2011, the best we’d produced was a ribozyme (called R18) that could polymerize just 14 ribonucleotides.  

Then a study was published showing that a modification of R18 could synthesize much longer strings and could replicate many different RNA templates. In this publication, the authors could synthesize ribonucleic acids of 95 bases, almost as long as the R18 replicase itself. Another study has shown that some catalytic RNAs can self-replicate at an exponential rate, making thousands of copies of themselves while still having catalytic function.

It seems that the RNA hypothesis is getting stronger, but there remain some hurdles.
A July, 2013 study shows that primitive protein enzymes (called urenzymes, where ur = primitive) activate tRNAs much faster than do ribozymes. These primitive proteins date to before the last common ancestor, so they have been around nearly as long as life itself. tRNA urenzymes suggest a tRNA-enzyme co-evolution, providing evidence that catalytic proteins and the conventional central dogma were important in early life – a result that does not support the RNA world hypothesis. I’m glad – the hunt goes on.

In the next weeks, let’s take a look at nucleic acid structures and their building blocks. Think DNA is double stranded? – not always. Think A, C, G, T, and U are the only nucleotides life uses? – not even close.

Chang W, Jääskeläinen M, Li SP, & Schulman AH (2013). BARE Retrotransposons Are Translated and Replicated via Distinct RNA Pools. PloS one, 8 (8) PMID: 23940808

Li L, Francklyn CS, & Carter CW (2013). Aminoacylating Urzymes Challenge the RNA World Hypothesis. The Journal of biological chemistry PMID: 23867455

Ferretti AC, & Joyce GF (2013). Kinetic properties of an RNA enzyme that undergoes self-sustained exponential amplification. Biochemistry, 52 (7), 1227-35 PMID: 23384307

For more information or classroom activities, see:

Nucleic acids –
Central dogma of molecular biology –

Types of RNA –

Retrotransposons –

RNA world hypothesis –

Catalytic RNA (ribozymes) –

Wednesday, September 18, 2013

It’s Not Just Our Tooth That’s Sweet

Biology concepts – homochirality, carbohydrates, chiral discrimination, glycoside, H antigen

It isn’t just biomolecules that show chirality. There is also
chiromorphology, like snail shells that usually turn to the right
(dextral, or D-). There are factors in early embryonic
development that cause the body and shell to be right handed
in most gastropod species, yet other species are left handed.
There are also instances where a right-handed species will
produce a left-handed individual, so shell collectors have to be
on the look out for abnormal individuals.
A couple of weeks ago we talked about how, in most cases, life uses exclusively the left-handed enantiomers of amino acids to make proteins.  This homochirality is also see in the sugars we talked about last week, but in this case, mostly D-sugars are utilized in biological systems.

What isn’t amazing is that it happens to be L- for amino acids and D- for carbohydrates; the fact that they’re different is no big deal. Evolution just wants the parts to fit together, so if an enzyme evolved to use D-sugars, it’s not a surprise that the D-sugar would be favored in the pathway then now on. 

But it might not have been random either. No one knows for sure, but hypotheses abound for how homochirality in these biomolecular monomers was established.

One 2009 paper was concerned with the maintenance of homochirality rather than its establishment. Dr. Soren Toxvaerd stated that if you don’t believe life as we see it today occurred in a singular event, then it must have developed over a long period of time. Evidence indicates that small changes in the self-assembly of biomolecules took place over at least thousands of years.

If life took a long time to develop, then prebiotic (before life) earth must have been fairly stable in terms of enantiomer concentrations. But we know that homochiral solutions will turn to racemic mixtures (containing both L- and D- enantiomers) in a short time, days for amino acids and just hours for sugars. So how could the environment have been stable enough for life to develop over time?

One possible hypothesis about the establishment of homochirality
was put forth in 2010 by Koji Tamura, PhD in the Journal of
Cosmology. Put very simply, RNA may have developed before
proteins. RNA evolved to use only D-ribose because a mixture
would have been a symmetry violation. The action of D-ribose
would have been driven toward L-amino acids because of shape
problems with attaching D-amino acids to tRNAs. Now prove it.
Louis Pasteur, he of bacteria-free milk and germ theory, may have shown us the way. He discovered chiral discrimination. Racemic mixtures, under the right conditions, will separate into pools of homochirality. There is an energy gain and stability to packing homochiral molecules together; the other enantiomer will be excluded. This could help explain life using one enantiomer only.

What is more, hydrothermal vents and black smokers have just the needed conditions for both chiral discrimination and for self-assembly of biomolecules. Interesting huh? Think it’s a coincidence that black smokers harbor some of the oldest archaea on Earth? We may owe our very existence to plumes of superheated water and the xenophobia of enantiomers.

Lastly in this area, it may be that sugars and amino acids selected each other for homochirality. Glyceraldehyde is 1) highly discriminate for its enantiomers, 2) was present in large amounts in prebiotic oceans, 3) is used in self-assembly of many biomolecules, and 4) D-glyceraldehyde very much likes to bind to L-serine. So a slight excess in either one of these could have helped select for the other, and if this was stable, it could have caught on like “Gangnam Style.” This may be why life uses mostly D-sugars and L-amino acids and why I know the name Psy.

Now that we have delved into the mire that is maintenance of homochirality in sugars, let’s look at the rule breakers. D-sugars aren’t the only game in town.

Bacteria, oh bacteria! Once again, they lead the way in rule breaking. Last week we discussed how E.coli can generate ATP from several different sugars - glucose, lactose, etc. It takes different enzymes to metabolize each sugar, so if they are going to invest the energy in maintaining those genes and making those enzymes, there better be a good reason.

Paracoccus species 43P  has been shown to have an L-glucose
metabolic pathway. This organisms is very closely related to
Paracoccus denitrificans. P. denitrificans is believed to be the
organism that was engulfed to become the eukaryotic
mitochondrion. It closely resembles the mitochondrion, and
although random genes needed for aerobic respiration have
been found in many prokaryotes, P. denitrificans is the only
prokaryote in which all the necessary genes have been found.
A 2012 study tried growing soil bacteria on medium that contained only L-glucose as an energy source. One species of bacterium, Paracoccus sp. 43P, was able to metabolize L-glucose to pyruvate and glyceraldehyde-3-P, and then make use that for ATP production. The researchers discovered an L-glucose-specific dehydrogenase enzyme, and this enzyme was active in the fluids from broken up paracoccus cells. The process is similar to one in E. coli, but here it is L-glucose specific.

Mammals can’t manage as well as some bacteria; we can’t metabolize L-glucose at all. However, that doesn’t mean it can't work for us. L-glucose has been proposed as an artificial sweetener, especially for type II diabetics. One form of L-glucose can stimulate insulin release, so this would be doubly good for type II diabetics. Unfortunately, L-glucose costs 50% more than gold; therefore, don't look for it next to the Truvia anytime soon.

One, but only one, study has been published showing rats metabolized L-fructose and L-gulose, but not L-glucose. From 1995, the authors waited until the end of the paper to explain that the metabolism was being carried out by the rodents gut bacteria, not by the rats themselves. No wonder it was only one paper.

Just because we can’t metabolize L-sugars doesn’t mean that we mammals are left out in the cold. Some sugars are used in the L-form even if they aren’t broken down to make ATP. The most egregious example of this is a hexose sugar called L-altrose. Why is it different than some other exceptions here? Because altrose doesn’t even occur in nature as a D-sugar; only the L-form has ever been found. It was first isolated in 1987 from a bacterium called, Butyrvibrio fibrisolvens, which is found in the GI tract of ruminate animals (cows and such).

Ruminants are mammals that have more involved digestive
strategies. Ruminants have many types of GI bacteria to help
them break down tough plant material; it isn’t surprising that
some of them can use nonstandard carbohydrates in their
physiology.  “Ruminating” is the act of re-chewing food that
has been partially softened by bacterial action in the first
compartment of the stomach, and then brought back to the
mouth as “cud.” I ruminate on ideas all the time, but I think I
will stop – I’m going to call it “further thought” from now on.
Ruminates go the extra mile. They digest longer and work on food harder, using bacteria to help with much of the work. Therefore, it isn’t strange to note that L-altrose has also been seen in another ruminate bacterium, Yersinia enterolitica. Remember though, this altrose isn’t being used in energy production; it's found in their outer cell wall glycoprotein, LPS (lipopolysaccharide).

It turns out that L-sugars are common in bacterial LPS. I found examples from several different bugs, including L-quinvose (6-deoxy-L- glucose), L-rhamnose, and L-fucose (6-deoxy–L- galactose).

When it comes to L-sugars, plants can get into the act as well. Rhamnose  (6-deoxy-L-mannose) occurs in nature, and can be isolated from several plants of the genera Rhamnus and Uncaria, including Buckthorn, poison sumac, and many other plants.

Rhamnose from plants takes the form of a glycoside. There’s there word again, glyco-. A glycoside in general terms is any molecule bound to a sugar. In plants, attaching sugars to create glycosides is a common way to inactivate molecules so that they can be stored for later use. When needed, the sugar residues of glycosides are cleaved away by special enzymes and then the protein, enzyme, lipid, etc. becomes active.

Digoxin (or sometimes digitalis) are cardiac glycosides from
foxglove plants. They are used to treat atrial rhythm or heart
failure problems. First used by William Withering in 1785,
digitalis is said to be the first of the modern day therapeutics.
But it can kill you too, both the plants and the drugs. A nurse
was sentenced to 18 life sentences after he was convicted of
killing more than 40 patients with digoxin.
Glycosides can be differentially regulated because there are many sugars that can be used, and several different possible linkages for each sugar/substrate combination. Therefore, cells can precisely control just when and where the glycosides are activated. This may allow cells to function for longer periods of time, but isn’t the reason that rhamnose and fucose (both L-sugars) are being included in obscenely expensive anti-aging creams.

Some evidence suggests that rhmanose and fucose can inhibit the activation of the elastase enzyme in skin cells. Elastase is known to increase in expression and activity as skin cells in culture divide several times. Therefore, companies want you to believe that rhamnose will keep your skin from looking old. Forget that keratinocytes in a petri dish bear as much resemblance to your skin as Watchmen does to Hamlet.

That was a bit sarcastic, but the cosmetic industry is a pet peeve of mine. And while I’m exposing my soul, I might as well admit to being a bit of a speciesist. I like the exceptions best when they involve Homo sapiens, so the last exception for today has to do with our own uses for a deoxy-L-sugar, fucose. I must admit that several uses of fucose apply to many mammals, but being the speciesist that you know I am, I ignore them to focus on humans.

Fucose (6-deoxy–L-galactose) is crucial for the turning of an unloved spermatozoa and a lonely oocyte into very premature teenager. Both the development and maturation of gamete cells and the development of the embryo depend on the recognition and communication of surface molecules that include fucose. But wait, there’s more.

The H antigen is linked to the red blood cell through a fucose
residue, but not in the “h” antigen mutant. Because of this, it
is not recognized for modification to the A or B antigen, and
the typical H antigen is not there to prevent development of
the H antibody.
Fucose is also a component of many glycans, including substance H. Also called the H antigen, this molecule is a precursor to the A and B antigens found on red blood cells. For people with A, B, or AB blood, the H antigen is modified to become the mature A or B antigen, but in people with O blood, the H antigen doesn’t mature and remains an H. Therefore, principal factors in every human’s development and physiology are determined in part by a sugar that we shouldn’t be using – according to the rules anyway.

However, not all is goodness and light when it comes to fucose. Some folks have a mutation in their H antigen gene that prevents its maturation to the A or B antigen. All cells would have the mutant H antigen, called h. This is different from being type O (meaning not having any A or B antigen, but still having the H antigen).

The hh or Oh blood type is called the Bombay type, and is very rare. Bombay individuals can donate blood to anyone, regardless of blood type (because they do not express any antigen to be attacked). However, because they make A, B, and H antibodies, they can receive blood only from another person with Bombay blood type. Since Bombay occurs about three times in a million births – good luck with that search for blood.

Let’s tackle the nucleic acids and their exceptions starting next week. By training I am a molecular biologist; I know an exceptional number of nucleic acid exceptions.

Shimizu T, Takaya N, & Nakamura A (2012). An L-glucose catabolic pathway in Paracoccus species 43P. The Journal of biological chemistry, 287 (48), 40448-56 PMID: 23038265

Toxvaerd S (2009). Origin of homochirality in biosystems. International journal of molecular sciences, 10 (3), 1290-9 PMID: 19399249
For more information or classroom activities, see:

Bombay blood type –

Glycosides –

Racemization –

Wednesday, September 11, 2013

Sugars Speak In Code

Biology concepts – carbohydrates, monosaccharides, hexose, glycocode, starch, glycogen, carbohydrate linkage, bacterial persisters, fructolysis

Refined sugar is produced from two main sources, sugar cane (37
different species of grass from the genus Saccharum, bottom right),
and sugar beet (Beta vulgaris, top right). Sugar cane accounts for
80% of the sugar produced today. The cane or the beets are ground
and the sugary juice is collected with water or on its own. To refine
the sugar, which still has molasses from the fiber, is processed with
lime or soda and evaporated to produce crystals. The color is
removed by activated charcoal to produce the white sugar we most
often see (top middle). Brown sugar is sugar in which the molasses
has not been removed and still coats the crystals (bottom middle).
Unprocessed sugar from cane is shown on the bottom right, while raw
sugar (not whitened) is on the top right.
It would be hard to argue that without sugars, none of us would be here. Glucose provides us with short and medium term storage of energy to do cellular work, but would you believe that certain parts of reproduction use a completely different energy source. All hail fructose!

Sugars are better termed carbohydrates, because they are basically carbon (carbo-) combined with water (-hydrate). The general formula is Cn(H2O)n; for instance, the formula for glucose is C6H12O6.

The simplest sugars are the monosaccharides (mono = one, and sacchar from the Greek = sugar. They can be composed of 4-7 carbons, called tetroses (4 carbon sugars), pentoses (5), hexoses (6), and septoses (7).

Things aren’t so simple though, even for the simple sugars. Let’s use the hexoses as an example, although what we say will also apply to the other sugars. We said the formula for glucose is C6H12O6, so that makes it a hexose. Is it the only hexose – heck no! Hexoses can be aldoses or ketoses, depending on their structure (see picture). Even more confusing, -OH groups can be located on different carbons making them act different chemically.

This chart is a brief introduction to the complexities of simple
sugars. They can vary in the number of carbons (triose vs.
pentose vs. hexose. They can also vary in their structure even
if they have same number of carbons (glucose vs. galactose).
Yet another difference can come in their reactive group on the
end, being either a ketone group (ketoses) or an aldehyde
group (aldoses).
There are actually 12 different hexoses – some names you know; glucose, fructose, or galactose. Others are less common; idose, tagatose, psicose, altrose, gulose – you won’t find those in your Twinkies. Then there are the deoxysugars, carbs that have lost an oxygen. Fucose is also called 6-deoxy-L-galactose, while 6-deoxy-L-mannose is better known as rhamnose.

If this wasn’t difficult enough, stereoisomers again rear their ugly head, as it did last week with the proteins. Hexoses have three (ketoses) or four (aldoses) chiral carbons each so hexoses can have eight or 16 stereoisomers! Every isomer may act differently from every other; this allows for many functions. But wait – there’s more trouble when we start linking sugars together.

Simple sugars can be joined together to build disaccharides (two sugars), oligosaccharides (3-10), and polysaccharides (more than 10). The subunits are connected by a hydrolysis reaction. Just like with the amino acid linkages in proteins, a water molecule is expelled when two sugars are joined together. Sucrose (table sugar) is a disaccharide made up of a glucose linked to a fructose.

Just where the linkage takes place is also important. Our example again can be glucose. Many glucoses can be linked together with an alpha-1,4 linkage. Long chains of glucoses linked in this way are called starch or glycogen, based on the different branching patterns they show. Mammals store glucoses as glycogen, while plants store them as starches.

Amylose is one type of starch, amylopectin being another.
They are different from celluloses only by the way the sugars
are linked together. You can see that in starch the CH2OH
group are all on the same side, while in cellulose they alternate.
This may seem like a small difference, but we can digest only
starch (or glycogen, which has the same type linkages),
not cellulose.
Humans can digest both starch and glycogen because we have enzymes that can break alpha-1,4 linkages. But if you change the chemical shape of the bond (see picture) to a beta-1,4 linkage, the glucose polymer becomes cellulose.

Plants make a lot of cellulose for structure, but even though it is made completely of glucose, humans can’t digest it at all! Ruminate animals can digest cellulose, but it takes some powerful gut bacteria to help out, and one of the side effects is a powerful dose of methane. Cows are the greatest source of methane on the planet!

We have talked about carbohydrates as energy sources, but pretty much every biological function and structure in every form of life involves carbohydrates.

Carbohydrates are important structural elements. Cellulose, thousands of beta-1,4-linked glucoses, help give plants their rigidity, especially in non-woody plants, but in woods as well (linked together by lignin). As such, cellulose is by far the most abundant biomolecule on planet Earth.

Chitin is another structural carbohydrate. Chitins make up the spongy material in mushrooms, and the crunchy stuff of insect exoskeletons.  You don’t get much more structural than keeping your insides inside.

Carbohydrates are often part of more complex molecules as well. Nucleic acids like RNA and DNA have a five-carbon ribose or deoxyribose at the core of their monomers. Glycolipids and glycoproteins (glyco- from Greek, also means sweet) are common in every cell. Over 60% of all mammalian proteins are bound to at least one sugar molecule.

The different sugar-linked complexes are part of the glycome (similar to genome or proteome), including oligo- and polysaccharides, glycoproteins, proteoglycans (a glycoprotein with many sugars added), glycolipids, and glycocalyxes (sugar coats on cell surfaces). None of these carbohydrate additions are coded for by the genetic code, yet a great diversity of glycomodifications are found on most structures of the cell.

The carbohydrate code is still a mystery to us. The glycosylation can be
linked together by N-type or O-type linkages, the order of the sugars
can vary, the numbers of each type of sugar can vary, and the branching
can vary. Every difference adds to the complexity of the code and can
direct a different message to the cell or the molecules with which
these glycans come into contact.
The diversity and complexity of these added carbohydrates is highly specific and highly regulated – this is the glycocode or carbohydrate code. Yet, we haven’t even come close to breaking the code, i.e., what series of what sugars means what.

The glycocode is important for cell-cell communication, immune recognition of self and non-self, and differentiation and maturation of specific cell types. Dysfunction in the glycocode leads to problems like muscular dystrophy, mental defects, and the metastasis of cancer – we better get cracking on the code breaking.

In the middle of 2013, a new method was developed for detecting the order and branching of sugars on different molecules. This method uses atomic force microscopy (AFM) to actually bump over the individual sugars on each molecule and identify them by their atoms, even on live cells. I’m proud to say that my father-in-law played a role in developing AFM for investigation of atom distributions on the surfaces of solid materials, mostly superconductors.

The glycome is even more diverse because different types organisms make different sugars. One thing I find interesting is that mammals don’t make sucrose. No matter what we mammals do, we won’t taste like table sugar when eaten – more’s the pity. I wonder what a sweet pork chop might taste like.

Proof that many foods have sugars – the Maillard reaction. That gorgeous
browning of your bread or steak comes from a chemical interaction
between the sugars and amino acids of the food. In the process, hundreds
of individual different compounds are made, each with a different flavor
profile. The example in the chart above is for caramelizing onions. Each
food and its chemical make up produces a different set of Maillard
products. You roast your coffee beans for the same reason. This is why
Food Network always suggests ways for you to get great searing and
browning of food.
We use sucrose as sugar because it is relatively easy to obtain from the plants that do make, like sugarcane or sugar beets. Fructose (often called fruit sugar) is actually sweeter on its own; almost twice as sweet as sucrose and three times as sweet as glucose.  This explains why so many sweetened foods are full of high fructose corn syrup (go here for our previous discussion of high fructose corn syrup).

We all know that organisms use glucose as an energy source, first through its breakdown to pyruvate via glyceraldehyde -3- phosphate (G3P) in glycolysis; the pyruvate then travels through the citric acid cycle to produce enough NADH and NADPH to generate a lot of ATP. But fructose can be used as well.

Fructose undergoes fructolysis, different from glycolysis only in the fact that one more step must be taken to generate G3P (adding the P to G3 is done by the enzyme trioskinase). In humans, almost all fructose metabolism takes place in the liver, as a way to either convert fructose to glucose to make glycogen, or to replenish triglyceride stores – so be good to your liver.

The big exception is how important fructose is in mammalian reproduction. Spermatozoa cells use fructose as their exclusive carbohydrate for production of ATP while stored in the testes. This fructose comes not from the diet but the conversion of glucose to fructose in the seminal vesicles.

Why use a different carbohydrate source just for sperm? Seminal fluid is high in fructose, not glucose. Perhaps this is a factor in seminal fluid viscosity. If this problem is solved using fructose, then the cells swimming in it would probably switch evolve to use it as an energy source.

I asked Dr. Fuller Bazer of Texas A&M about this and he pointed out that fructose can be metabolized several different ways, and some of these lead to more antioxidants and fewer reactive oxygen species - it would be important to leave sperm DNA undamaged, especially since we have previously talked about how they are more susceptible to oxidative damage.

Bazer also pointed out that unlike glucose, fructose is not retrieved from tissues and put back into circulation. Once it’s sequestered to the male sexual accessory glands, it would stay there. Still lots to be learned in this area.

Fructose is sweeter than glucose. Sucrose is one glucose joined to one
fructose, so the ratio is 50:50. In most honey, the fructose:glucose ratio
is about 55:45, so it is often sweeter than table sugar. Since it is higher
in fructose, some people liken it to high fructose corn syrup, but there
are many compounds in honey that also help the immune system, etc.
However, recent evidence is showing that some honey is being diluted
with high fructose corn syrup and some bees are being fed HFCS. The
benefits from true honey are then lost.
A 2013 study shows that maternal intake of fructose can also affect reproduction. Pregnant rats fed 10% fructose in their drinking water had significantly fewer babies, but a greater percentage of the offspring were male (60% versus 50%). The fructose did not arrest female embryos from developing or have a sex-specific effect on sperm motility, suggesting that the sugar has a direct effect on the oocyte that increases the chances of being fertilized to produce a male. Weird.

Using sugars other than glucose may be a big deal for mammals, but bacteria can thrive on many different sugars. E. coli can process glucose, but if other sources of sugar are around, they will switch over in a heartbeat – if they had a heart. E. coli has a whole different set of genes for lactose metabolism, found in something called the Lac operon. The operon gets turned on only if lactose is present and glucose is not.

The ability for bacteria to use other sugars might save us as well. Some bacteria can just shut down their metabolism if antibiotics are present and just hangout until the drugs are gone. These are called persister organisms, and they are different from antibiotic resistant bacteria. A 2011 study showed that if you give sugar in combination with some kinds of antibiotics, the persisters just can’t resist the sweet treat and will not shut down their metabolism. The antibiotics then become effective. Using sugars we don't metabolize, like fructose or mannitol, ensures that they will be around to help kill the bacteria. Amazing.

We have just brushed the surface of sugary exceptions. Next week we will see how nature first selected a single type of sugar to use in biology, and then went right out and broke its own rule.

Gunning AP, Kirby AR, Fuell C, Pin C, Tailford LE, & Juge N (2013). Mining the "glycocode"--exploring the spatial distribution of glycans in gastrointestinal mucin using force spectroscopy. FASEB journal : official publication of the Federation of American Societies for Experimental Biology, 27 (6), 2342-54 PMID: 23493619

Gray C, Long S, Green C, Gardiner SM, Craigon J, & Gardner DS (2013). Maternal Fructose and/or Salt Intake and Reproductive Outcome in the Rat: Effects on Growth, Fertility, Sex Ratio, and Birth Order. Biology of reproduction PMID: 23759309

Allison KR, Brynildsen MP, & Collins JJ (2011). Metabolite-enabled eradication of bacterial persisters by aminoglycosides. Nature, 473 (7346), 216-20 PMID: 21562562  

For more information or classroom activities, see:

Testing for carbohydrates in foods –

Structures of carbohydrates –

Glycocode/carbohydrate code –