You Gotta See This ...

 
Jason Rosenhouse says you gotta see this. I agree. It's how I feel every time I try and use Adobe Photoshop.

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The Peace Tower Clock Does Not Fall Back Tonight

 
Canada likes to think of itself as a progressive country—always moving forward. But this seems to be going to extremes. The clock on the Peace Tower (Parliament Buildings) does not go backwards. Thus, according to CBCNews [Time stops annually on Parliament Hill as Peace Tower clock falls back]...

While most Canadians scurry around their homes changing their clocks back to standard time this weekend, the clock-keepers of Parliament Hill will only sit and wait.

At 2 a.m. Eastern Daylight Time on Sunday, a Public Works employee will open a glass housing and flip a switch, bringing the 50-year-old mechanism that runs the Peace Tower clock, with its four faces, to a halt....

For 60 minutes, the technician will just wait and then will restart the 1950s-era electric motor drive that runs the big clock, several levels overhead, at exactly 2 a.m. Eastern Standard Time, according to the National Research Council time signal

They need to wait out the hour because the old motor drive only goes forward, says Brian Cook, the Public Works property manager for Parliament Hill.

At the risk of sounding stupid, why doesn't the technician just advance the clock eleven hours?

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You poor Amercans ....

 
I love the wit and humor of Canadian Cynic. From time to time he comes up with some very funny lines. Today is one of those times [ Suddenly, the parallels are starting to creep me out].

You poor Americans -- you have a leader whose party doesn't even have a majority, who's submitting one vile bit of legislation and one horrendous nomination after another, and you have an "opposition" that just rolls over and plays dead on all of it. I feel so sorry for you. Now up here in Canada, we ... we ... um ... actually, never mind.

I should have thought about that argument a bit harder, really.

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Dawkins on Watson

 
There's a lengthy article on the Guardian website about the Watson affair [Disgrace: How a giant of science was brought low]. It contains quotations from Richard Dawkins and Oxford neurologist Colin Blakemore,

In the end, Watson's decided to return home, so no meetings occurred, a move that has dismayed many scientists who believed that it was vital Watson confront his critics and his public. 'What is ethically wrong is the hounding, by what can only be described as an illiberal and intolerant "thought police", of one of the most distinguished scientists of our time, out of the Science Museum, and maybe out of the laboratory that he has devoted much of his life to, building up a world-class reputation,' said Richard Dawkins, who been due to conduct a public interview with Watson this week in Oxford.

Dawkins's stance was supported by Blakemore. 'Jim Watson is well known for being provocative and politically incorrect. But it would be a sad world if such a distinguished scientist was silenced because of his more unpalatable views.'

I agree with Dawkins. Watson was stupid to make those remarks but they were perfectly consistent with a lifelong career of being as politically incorrect as possible in today's society. Does that make him a racist whose career should be terminated?

Nor is it at all clear that Watson is a racist, a point stressed last week by the Pulitzer-winning biologist E O Wilson, of Harvard University. In his autobiography, Naturalist, Wilson originally described Watson, fresh from his Nobel success, arriving at Harvard's biology department and 'radiating contempt' for the rest of the staff. He was 'the most unpleasant human being I had ever met,' Wilson recalled. 'Having risen to fame at an early age, [he] became the Caligula of biology. He was given licence to say anything that came into his mind and expected to be taken seriously. And unfortunately he did so, with casual and brutal offhandedness.'

That is a fairly grim description, to say the least. However, there is a twist. There has been a rapprochement. 'We have become firm friends,' Wilson told The Observer last week. 'Today we are the two grand old men of biology in America and get on really well. I certainly don't see him as a Caligula figure any more. I have come to see him as a very intelligent, straight, honest individual. Of course, he would never get a job as a diplomat in the State Department. He is just too outspoken. But one thing I am absolutely sure of is that he is not a racist. I am shocked at what has happened to him.'

This is a clear case of political correctness out of control. I'm embarrassed to be associated with the people who attacked Watson and I admire Dawkins (and Blakemore) for standing up to them.

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Berlinski Quotes

 
In the comments section of Breaking News... Mathematicians Don't Believe in Evolution!, glennd points us to An Interview with David Berlinski on an Intelligent Design Creationist website. (Incidentally, the commenters on that thread do a good job of proving that Berlinski isn't a mathematician.

Here's are some of the eye-popping views of this "famous" mathematician "philosopher" ...

The Panda’s Thumb, on the other hand, is entirely low-market; the men who contribute to the blog all have some vague technical background – computer sales, sound mixing, low-level programming, print-shops or copy centers; they are semi-literate; their posts convey that characteristic combination of pustules and gonorrhea that one would otherwise associate with high-school toughs, with even the names – Sir Toejam, The Reverend Lenny Flank – suggesting nothing so much as a group of guys spending a great deal of time hanging around their basements running video games, eating pizzas, and jeering at various leggy but inaccessible young women.

Darwin’s theory is plain nuts. It is not supported by the evidence; it has no organizing principles; it is incoherent on its face; it flies against all common experience, and it is poisonous in its implications.

And another thing. It is easy to understand. Anyone can become an evolutionary biologist in an afternoon. Just read a book. Most of them are half illustrations anyway. It’s not like studying mathematics or physics, lot of head splitting stuff there.

But Dawkins …

DB: An interesting case, very louche – fascinating and repellant. Fascinating because like Noam Chomsky he has the strange power effortlessly to command attention. Just possibly both men are descended from a line of simian carnival barkers, great apes who adventitiously found employment at a circus. I really should look at this more closely. Repellent because Dawkins is that depressingly familiar figure – the intellectual fanatic. What is it that he has said? “It is absolutely safe to say that, if you meet somebody who claims not to believe in evolution, that person is ignorant, stupid or insane (or wicked, but I’d rather not consider that)”. Substitute ‘Allah’ for ‘evolution,’ and these words might have been uttered by some fanatical Mullah just itching to get busy with a little head-chopping. If he ever gets tired of Oxford, Dawkins could probably find a home at Finsbury Park.

It is a matter of attitude and sentiment, Look, for thousands of intellectuals, becoming a Marxist was an experience of disturbing intensity. The decision having been made, the world became simpler, brighter, cleaner, clearer. A number of contemporary intellectuals react in the same way when it comes to the Old Boy – Darwin, I mean. Having renounced Freud and all his wiles, the literary critic Frederick Crews – a man of some taste and sophistication – has recently reported seeing in random variations and natural selection the same light he once saw in castration anxiety or penis envy. He has accordingly immersed himself in the emollient of his own enthusiasm. Every now and then he contributes an essay to The New York Review of Books revealing that his ignorance of any conceivable scientific issue has not been an impediment to his satisfaction.

Another example – I’ve got hundreds. Daniel Dennett has in Darwin’s Dangerous Idea written about natural selection as the single greatest idea in human intellectual history. Anyone reading Dennett understands, of course, that his acquaintance with great ideas has been remarkably fastidious. Mais, je divague …

In the case of both Crews and Dennett, it’s that God-awful eagerness to explain everything that is the give-away. The eagerness is entirely academic or even literary. But, you know, what sociologists call prole-drift is present even in a world without proles. Look at Christopher Hitchens – very bright, very able. Just recently he felt compelled to release his views on evolution to a public not known eagerly to be waiting for them. What does he have to say? Pretty much that he doesn’t know anything about art but he knows what he likes. The truth of the matter, however, is that he pretty much likes what he knows, and what he knows is what he has heard smart scientists say. Were smart scientists to say that a form of yeast is intermediate between the great apes and human beings, Hitchens would, no doubt, conceive an increased respect for yeast. But that’s a journalist for you: all zeal and no content. No, no, not you, of course. You’re not like the others.

The problem with Berlinski is that he's a fuzzy-headed idiot on some things but on others he hits a little too close to the mark for my comfort level. He's the Christopher Hitchens of the Intellligent Design movement.

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[Photo Credit: Turkey's First ID Conference, March 2007]

Chalk Up One for the Intelligent Design Creationists

 
Jason Rosenhouse has been following an exchange between Micheal Behe and Theistic Evolutionist Creationist Ken Miller [In Which I Agree With Michael Behe Over Ken Miller]. Miller is upset because Behe's intelligent designer made malaria.

Miller doesn't think this fits with his idea of a loving God so he criticizes Behe in a recent review published in a Catholic magazine. Behe responds and here's what Jason says,

Bingo! That's exactly right, and it nicely punctures the sophistry offered up by theistic evolutionists.

I agree with Jason. This is a fight between two Intelligent Design Creationists even though one of them (Ken Miller) pretends that he's not a creationist. What I like about this exchange is that Behe is honest and forthright about the implications of his belief in a designer. The Theistic Evolutionist Creationists on the other hand, aren't.

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Best Science Blog

 
There are ten candidates in the voting for Best Science Blog—part of the 2007weblogawards.

I'm familiar with two of them; Pharyngula and Bad Astronomy but not with the others. Can anyone help out? What's interesting in that list?

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Breaking News... Mathematicians Don't Believe in Evolution!

 

This jerk is David Berlinski a mathematician and a Fellow of the Discovery Institute's Center for Science and Culture. He's not a biologist and neither are his "skeptical" mathematician friends. I wonder what he would say if a bunch of evolutionary biologists expressed skepticism about vector calculus and scaler fields? They sound like pretty crazy ideas to me.

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Denyse O'Leary's Advice to Students

 
Denyse O'Leary is teaching a short course on intelligent design creationism [Denyse O'Leary's University Course on Intelligent Design]. Presumably, the students in her class don't need to be afraid of making fools of themselves but it's a different story when creationists sit in on a real science course.

Not to worry. She has an answer for those students [Not a Darwinbot? Got a story? Tell it to The EXPELLED!].

I usually tell students, keep QUIET. Act like a good little Darwinbot. Question nothing, no matter how ridiculous. Practice keeping a straight face. (Anyone who laughs will be disqualified.) Get great marks.

It's called lying for Jesus.

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Is Race a Biological Concept?

 
I suppose it was inevitable. The latest issue of New Scientist has the obligatory article denying that intelligence can be defined and denying that humans can be separated into races. This is required political correctness in light of Jim Watson's comments from two weeks ago.

The politically correct author in this case is Robert J. Sternberg, a psychologist at Tufts University [Race and intelligence: Not a case of black and white]. He writes ...

A further hugely complicating factor is what we mean by the word "race". Populations in different parts of the world have clearly adapted to their environments in different ways. A trait that is beneficial in one environment may work against people in another. Obesity is a problem today because it once was beneficial to eat as much as one could while one could. Stratification - classifying people into categories of higher and lower status in a society - already occurs on the basis of weight just as it has on the basis of intelligence test scores.

But there is nothing special about skin colour that serves as a basis for differentiating humans into so-called races. Skin colour correlates only weakly with genetic differentiations. Sarah Tishkoff, a geneticist at the University of Maryland, and Kidd have found that the genetic differences among black Africans are often greater than those between blacks and whites. The significance of those labels stems only from the fact that society has found it convenient to label races on the basis of skin colour.

Curiously, we do not apply the concept of "race" to colours of dogs or cats - or moths, for that matter. For some of these, colour can be important: being a black moth confers camouflage advantages in polluted environments and disadvantages in clean environments - and vice versa for white moths. Similarly, our ancestors in Africa were almost certainly dark-skinned because it provided better protection against the particular challenges of the environment, such as ultraviolet light. We could of course refer to moths as being of different "races". We do not, presumably because we are less interested in creating social classes for moths than for people.

The problems with our understanding of intelligence and race show that the criticism being levelled at Watson is based on science rather than political correctness. Intelligence is clearly a far more complicated issue than standard testing allows. And race is a socially constructed concept, not a biological one. It derives from people's desire to classify. Whether people with a genetic predisposition toward fatness will be classified as a separate race remains to be seen.

We all know what people mean when they talk about blacks and whites. Those are synonyms for Africans and Europeans. Unless Sternberg is being extremely pedantic, he's arguing that there are no such thing as distinct populations of Europeans and Africans that differ genetically. Races—or demes if you wish—don't exist according to him.

That's nonsense, of course, but it seems to be widespread nonsense. I'm beginning to wonder whether the discipline of psychology deserves to be called a science.

There's an interesting press release out today from Cold Spring Harbor Press [Scientists discover genetic variant associated with prostate cancer in African Americans]. It reports on a study of higher incidence of prostate cancer among African Americans compared to European Americans. The scientists identified a particular locus on chromosome 8 (8q24) that may contain a genetic variant that differs between the two groups.

Other studies show that the incidence of cystic fibrosis is higher among Europeans (whites) than among Africans (blacks).

How could there be a genetic difference between Africans and Europeans if there's no such thing as race? If these are just social constructs there shouldn't be any genetic differences that correlate with other features used to distinguish the two groups, right?

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What's Your Image?

 
PZ Myers is bored in San Diego so he came up with another meme for bloggers. This time we're supposed to Goggle for the first image that comes up when you enter your name [What's your image?].

Being a sucker, I fell for it. Guess what's the first photograph of real people that comes up with "Larry Moran"?

Right, it's a picture of PZ Myers (left) [A Visit to Downe]. How evil is that? Am I the only one this happened to? Is Pharyngula taking over the world?

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Can You Smell Isovaleric Acid?

 
Isovaleric acid [3-Methylbutanoic acid] smells like sweat. It is responsible for some of the odor in a locker room, for example. Although we can all detect that odor, some of us are much more sensitive to it than others. In fact, the concentrations of isovaleric acid that can be detected differs by as much as 10,000-fold from one individual to the next.

It turns out that the ability to detect the molecule has a genetic component. It's quite likely that many people reading this blog can't smell isovaleric acid at low concentrations because they don't have one of the olfactory receptors for that ligand [A Sense of Smell: Olfactory Receptors].

Mice have about 1000 genes for olfactory receptors and this single gene family accounts for about 4% of all the genes in the mouse genome. Since each receptor is presumably capable of binding a specific odorant, it seems very likely that mice can detect a large number of different smells.

Humans have about 800 olfactory receptor genes but half of them are pseudogenes. They are incapable of producing a full-length functional receptor protein. Thus, it is reasonable to conclude that humans can detect far fewer smells than mice can.

These conclusions are based on the assumption that each olfactory receptor can bind to a single odorant molecule—or a small number of related molecules. If this assumption is correct then it should be possible to identify specific olfactory receptor genes that are responsible for the diversity in odor detection. Menashe et al. (2007) decided to test this by surveying 377 individuals for their ability to detect four odorants: isoamyl acetate, isovaleric acid, L-carvone, and cineole. The authors then tried to correlate ability to detect low levels of these odorants with the presence of specific markers for alleles of olfactory receptor genes.

Theme
A Sense of Smell
There was a strong association between ability to detect low levels of isovaleric acid and an allele for OR gene OR11H7P. This particular allele (OR11H7Pi) is an active form of the gene whereas the other allele is a pseudogene. People who were homozygous for the pseudogene were much less sensitive to isovaleric acid whereas people who had one or two copies of the active gene could detect low levels of isovaleric acid.

It looks like OR11H7P encodes a receptor that binds isovaleric acid. In order to test this Menashe et al. cloned the active gene and inserted it into a frog oocyte detection system. The olfactory receptor encoded by this gene responded to isovaleric acid whereas the pseudogene produced no response and other intact genes did not respond to this ligand.


The OR11H7P gene is part of a large cluster of olfactory receptor genes on chromosome 14. (OR11H7P is the yellow triangle marked by two asterisks.) The two flanking genes (OR11H4 and OR11H6) are closely related to OR11H7P indicating recent duplication events. Menashe et al. also cloned and tested these genes in the in vitro assay and they responded to isovaleric acid as well. This probably explains the detectability of isovaleric acid in those people who lack a functional copy of OR11H7P.

The results demonstrate a direct link between phenotpypic variation in human olfaction and olfactory receptor gene polymorphisms. This linkage does not account for all of the variation in ability to detect isovaleric acid. In fact, the authors estimate that it accounts for less than 10% of the variation. The rest is probably due to polymorphisms or environmental differences in downstream parts of the olfactory detection pathway.

The results also show that there is a certain amount of redundancy in ligand binding to receptors. Closely related olfactory receptors molecules tend to bind similar odorants. The more kinds of active receptors present in the sensory neurons of the nasal cavity, the greater the capacity to detect low concentrations of odorant.

The authors note that the OR11H7P gene is identified as a pseudogene in the public databases [EntrezGene 390441]. The fact that they were able to discover a minor active allele is a warning to not assume that all annotated pseudogenes are necessarily inactive in all individuals.

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Menashe, I., Abaffy, T., Hasin, Y., Goshen, S., Yahalom, V., Luetje, C.W. and Lancet, D. (2007) Genetic Elucidation of Human Hyperosmia to Isovaleric Acid. PLoS Biology 5:e284 doi:10.1371/journal.pbio.0050284. [PLoS Biology]

Theme: A Sense of Smell

 
The detection of odor is a complex signal transduction pathway that begins with the binding of an odor molecule (ligand) to an olfactory receptor located in sensory neurons in the nasal cavity. The pathway is interesting for a number of reasons including the mechanism of signal transduction and the structure of the olfactory receptors. One of the important problems in the field is the identification of the specific odor molecules that bind to specific receptors—or even whether each receptor actually binds a specific molecule.

The olfactory receptor genes make up the largest gene family in mammalian genomes and the study of these genes and their evolution provides plenty of opportunities to learn about the mechanisms of gene family evolution.

Jan. 8, 2007
Monday's Molecule #8

Jan. 9, 2007
The Smell of Cat Pee

Jan. 9, 2007
A Sense of Smell: Olfactory Receptors

Jan. 10, 2007
Nobel Laureates: Richard Axel and Linda B. Buck


Jan. 11, 2007
Olfactory Receptor Genes

Jan. 13, 2007
The Evolution of Gene Families (Birth and Death)

Sept. 20, 2007
Calling All Adaptationists (Again)

Nov. 1, 2007
Can You Smell Isovaleric Acid?

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Nobel Laureate: Arthur Kornberg

 

The Nobel Prize in Physiology or Medicine 1959.

"for their discovery of the mechanisms in the biological synthesis of ribonucleic acid and deoxyribonucleic acid"

Arthur Kornberg (1918-2007) received the Nobel Prize in Physiology or Medicine for discovering an enzyme that replicates DNA. The enzyme, now known as DNA polymerase I, plays a key role in joining Okazaki fragments on the lagging strand and in DNA repair [DNA Polymerase I and the Synthesis of Okazaki Fragments] [Biochemist Arthur Kornberg (1918 - 2007)]. Kornberg shared the prize with Severo Ochoa.

The presentation speech was delivered by by Professor H. Theorell of the Royal Caroline Institute.

In the lessons I have learned from the enzymology of DNA replication, I depended at every turn on colleagues near and far for orientation and guidance. Most of all, I learned from the efforts and contributions of my students, too numerous to be mentioned individually. Without them there would be no story for me to tell.

Arthur Kornberg (2000)

Your Majesties, Your Royal Highnesses, Ladies and Gentlemen.

To maa man vaere hvis livet skal lykkes, «There must be two if life shall succeed», is the theme of a sentimental old Danish song. The author had in mind man and woman, but she probably did not know how right she was from a more elementary biological viewpoint. Two principles are necessary so that «life» shall «succeed». One consists of proteins, the other of nucleic acids. The analogy is more than just a play upon words. Just as man and woman are responsible for the regeneration of mankind, likewise is the interplay between proteins and nucleic acids the only, and universally repeated basic mechanism of life. In the long series of substances which build up viruses, bacteria, plants, and animals, everything else might vary, but the proteins and nucleic acids are always present as the life-supporting elements. These both show certain principal characteristics. Their molecules are very large, and are built up from thousands of smaller units linked together in chains - just like a string of pearls - which have a tendency to form helices. Single helices join together in complicated threads which can contain proteins or nucleic acids or both. In the mixed «super molecules», the reactions of life proceed in the subtle pattern of the intimately associated strands.

The proteins contain amino acids as their elementary part. In the whole of Nature on this earth, there are found only some twenty amino acids in proteins. The elementary parts of the nucleic acids, the nucleotides, consist of nitrogenous bases, sugar, and phosphoric acid. There are found in Nature practically no more than eight of these most important nucleotides, all of which contain phosphoric acid, but in which the nitrogenous base may be one of five different kinds. The sugar can be of two different kinds, one of which, «ribose», contains one more oxygen atom than the other, «deoxyribose». This seemingly insignificant difference in a single atom produces a remarkably great effect. Nucleic acids are divided into two different series because of this characteristic. These series have widely different functions, so widely different in fact, that this is the reason why we have two Prize Winners on the stage today.

«Deoxyribonucleic acids», which Arthur Kornberg has now synthesized, are mainly present as the hereditary substance in chromosomes. The «ribonucleic acids», which Severo Ochoa has synthesized, have other functions, such as to assist in the synthesis of proteins. The Swedish scientist Torbjörn Caspersson has played an important role in demonstrating this last fact. From his and other research-workers' discoveries, it has been possible to conclude that nucleic acids assist in the synthesis of protein. The exact chemical mechanism, however, is as yet unknown. Inasmuch as nucleic acids and proteins are the two main principles of life, it seemed highly probable that, vice versa, the proteins should take part in the rebuilding of nucleic acids. This seems so much the more probable when we realize that proteins, in the form of enzymes, take part in practically every chemical reaction in the biological world. It is to the everlasting credit of Ochoa and Kornberg to have clarified this fundamental mechanism by preparing proteins that build up nucleic acids in test tubes.

For proteins it has been proved, and for the nucleic acids it is highly probable, that the order of the different building blocks in the chains is by no means left to chance, but on the contrary is planned in detail for each kind of molecule and for each kind of living organism.

It is this regulated order between the building blocks that always makes human children grow up to be human beings, and the serpent's offspring grow up to be serpents. It is disturbances in this regulated order which change the hereditary factors and allows the variations of species over thousands of years. The almost infinite possibilities to combine the building blocks in different ways makes it possible to vary the form in which life appears on our earth. Let me give a comparable example. By different combinations of the 28 letters in our alphabet, we can write everything that can be expressed in our own language, as well as in all other languages. The building blocks of the proteins, the amino acids, are approximately equal in number to the letters of the alphabet. The protein molecules can be compared with words with 100, 1,000, or even 10,000 letters. It is clear that Nature here has been generous with the possibility to make different combinations amounting to astronomical figures. But here, another factor might be brought up. The differences between the amino acids are necessary not only to produce the possibilities for variation, but also to enable the proteins, by their enzymatic activity, to regulate the different aspects of metabolism. Even the two types of nucleic acids with 4 different nucleotides in each, when made up from 100 to 10,000 nucleotide units in each molecule, give a fantastic number of possible combinations. Thus it would seem as if it were too heroic an enterprise to try to find out the procedure whereby Nature forms such complicated substances as nucleic acids with such an unerring accuracy in placing each building block.

A few years ago, Ochoa and Kornberg, each in his own laboratory, started to investigate the problem. The development turned for Ochoa's part in a direction that made him work with systems that produced ribonucleic acids, while for Kornberg it led him to investigate the formation of deoxyribonucleic acids. They have both, in a series of outstanding investigations, without direct cooperation, but nevertheless, as personal friends probably profiting from each other's results, reached the goal at the same time. As everyone else, they were able to make use of results obtained by preceding research workers, among whom I can only mention a few. It might be of interest to mention that uric acid, the first representative of the purines (a class of nitrogenous bases that form part of the nucleic acids) was discovered in Sweden in 1776 simultaneously by both Carl Wilhelm Scheele and Torbern Bergman. It is a curious parallel to the shared Nobel Award of today, and a reminder of Sweden's great era in the science of chemistry. The German scientist Albrecht Kossel received the Nobel Prize in 1910 for his elucidation of the chemistry of the nitrogenous bases of the nucleic acids, whereas the English scientist Alexander Todd clarified in detail the chemical properties of the nucleic acids, and received the Nobel Prize for Chemistry in 1957. But what really enabled Ochoa and Kornberg to succeed was their own excellent previous work in related fields. Both have worked with bacteria from which they have made preparations of a high degree of purity, Ochoa from an acetic acid bacterium, and Kornberg from the coli bacterium. Ochoa's enzyme produces ribonucleic acids from ribonucleotides having twice the ratio of phosphoric acid residues as that contained in ribonucleic acid. The ribonucleic acid is formed by splitting out half of the phosphoric acid residues, and linking the nucleotides together to form large molecules, which, as far as we can prove today, do not differ in any way from natural nucleic acids. Kornberg's enzyme produces deoxyribonucleic acids in a similar, but not identical fashion. Both have arrived at the same, principally important result that in order to make the reaction start, it is necessary to add in the beginning a small amount of nucleic acid to act as a template. Otherwise the enzymes do not «know» which kind of nucleic acid they are to produce. As soon as they get a template to act as a guide, they start, just like a skilled type-setter, to copy the «manuscript» they have received. Here one recognizes life's own principle that like creates like. Even though several research workers had earlier suspected that such a mechanism was involved, the actual experimental proof is of greatest importance. Furthermore, Ochoa's enzyme has given us the possibility of enzymatically synthesizing simplified nucleic acids of great interest.

To give an idea of where the discoveries that are being honored today may lead to in the near future, I want to mention one example. Other scientists, especially S. S. Cohen in the U.S.A., have shown that the nucleic acid of a certain bacteriophage, T2, which is a kind of bacterial virus, contains a somewhat chemically different nitrogenous base. If bacteria are infected with T2 phage, this different nucleic acid is soon produced in the bacteria. Kornberg succeeded in explaining the mechanism in detail. T2 phage behaves like the worst kind of usurper. Within four minutes it produces a number of enzymes which destroy a nucleotide necessary for the bacterium's normal production of nucleic acids, and rebuilds it to the different nucleotide of the T2 phage, and thereby destroys the bacteria.

We are sure to witness in the near future several important discoveries in biochemistry, virus research, genetics, and cancer research as a consequence of the work of Ochoa and Kornberg. They have helped us to advance quite some distance on the road to understanding the mechanism of life.

Professor Severo Ochoa, Professor Arthur Kornberg, dear friends and colleagues. Some 130 years ago, Friedrich Wöhler, in the laboratory of Berzelius, synthesized urea from inorganic matter. This event occurred in the heart of this city of Stockholm, less than half a mile from where we are now standing. He thus overbridged the first gap between living and dead material. You have now made the second fundamental discovery on this pathway, the synthesis in test tubes of one of the two basic principles of life.

On behalf of the Caroline Institute, I extend to you our warm congratulations, and ask you to receive this year's Nobel Prize for Physiology or Medicine from the hands of His Majesty the King.

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DNA Polymerase I and the Synthesis of Okazaki Fragments

 
Slightly modified from Horton et al. (2006) ...

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During DNA replication, a molecular machine called a replisome forms at the replication fork where the two strands of DNA are separating. The replisome contains activities that separate the strands and hold them apart for synthesis by the replisome version of DNA polymerase, called DNA polymerase III in bacteria. The complex has two sliding clamps that bind the complex to the strands of DNA so that DNA replication is highly processive.

DNA polymerases catalyze chain elongation exclusively in the 5′ → 3′ direction. Since the two strands of DNA are antiparallel, synthesis using one template strand occurs in the same direction as fork movement, but synthesis using the other template strand occurs in the direction opposite fork movement. The new strand formed by polymerization in the same direction as fork movement is called the leading strand. The new strand formed by polymerization in the opposite direction is called the lagging strand.

THEME

Deoxyribonucleic Acid (DNA)Recall that the replisome contains a DNA polymerase III holoenzyme dimer with two core complexes that can catalyze polymerization. One of these is responsible for synthesis of the leading strand, and the other is responsible for synthesis of the lagging strand.

Here's a video showing the entire process (source unknown, please contact me if you know who made this video). The details of one of the important steps are presented below the fold.



A. Lagging-Strand Synthesis Is Discontinuous

The leading strand is synthesized as one continuous polynucleotide, beginning at the origin and ending at the termination site. In contrast, the lagging strand is synthesized discontinuously in short pieces in the direction opposite fork movement. These pieces of lagging strand are then joined by a separate reaction.

The short pieces of lagging-strand DNA are named Okazaki fragments in honor of their discoverer, Reiji Okazaki. The overall mechanism of DNA replication is called semidiscontinuous to emphasize the different mechanisms for replicating each strand.

B. Each Okazaki Fragment Begins with an RNA Primer

It is clear that lagging-strand synthesis is discontinuous, but it is not obvious how synthesis of each Okazaki fragment is initiated. The problem is that no DNA polymerase can begin polymerization de novo; it can only add nucleotides to existing polymers. This limitation presents no difficulty for leading-strand synthesis since once DNA synthesis is under way nucleotides are continuously added to a growing chain. However, on the lagging strand, the synthesis of each Okazaki fragment requires a new initiation event. This is accomplished by making short pieces of RNA at the replication fork. These RNA primers are complementary to the lagging strand template. Each primer is extended from its 3′ end by DNA polymerase I to form an Okazaki fragment, as shown in the Figure. (Synthesis of the leading strand also begins with an RNA primer, but only one primer is required to initiate synthesis of the entire strand.)

The use of short RNA primers gets around the limitation imposed by the mechanism of DNA polymerase, namely, that it cannot initiate DNA synthesis de novo. The primers are synthesized by a DNA dependent RNA polymerase enzyme called primase—the product of the dnaG gene in E. coli. The three-dimensional crystal structure of the DnaG catalytic domain revealed its folding and active site are distinct from the well studied polymerases, suggesting that it may employ a novel enzyme mechanism. Primase is part of a larger complex called the primosome that contains many other polypeptides in addition to primase. The primosome, along with DNA polymerase III, is part of the replisome.

As the replication fork progresses, the parental DNA is unwound, and more and more single-stranded DNA becomes exposed. About once every second, primase catalyzes the synthesis of a short RNA primer using this single-stranded DNA as a template. The primers are only a few nucleotides in length. Since the replication fork advances at a rate of about 1000 nucleotides per second, one primer is synthesized for approximately every 1000 nucleotides that are incorporated. DNA polymerase III catalyzes synthesis of DNA in the 5′ → 3′ direction by extending each short RNA primer.

C. Okazaki Fragments Are Joined by the Action of DNA Polymerase I and DNA Ligase

Okazaki fragments are eventually joined to produce a continuous strand of DNA. The reaction proceeds in three steps: removal of the RNA primer, synthesis of replacement DNA, and sealing of the adjacent DNA fragments. The steps are carried out by the combined action of DNA polymerase I and DNA ligase.

DNA polymerase I of E. coli was discovered by Arthur Kornberg about 45 years ago. It was the first enzyme to be found that could catalyze DNA synthesis using a template strand. In a single polypeptide, DNA polymerase I contains the activities found in the DNA polymerase III holoenzyme: 5′ → 3′ polymerase activity and 3′ → 5′ proofreading exonuclease activity. In addition, DNA polymerase I has 5′ → 3′ exonuclease activity, an activity not found in DNA polymerase III.

DNA polymerase I can be cleaved with certain proteolytic enzymes to generate a small fragment that contains the 5′ → 3′ exonuclease activity and a larger fragment that retains the polymerization and proofreading activities. The larger fragment consists of the C-terminal 605 amino acid residues, and the smaller fragment contains the remaining N-terminal 323 residues. The large fragment, known as the Klenow fragment, is widely used for DNA sequencing and many other techniques that require DNA synthesis without 5′ → 3′ degradation. In addition, many studies of the mechanisms of DNA synthesis and proofreading use the Klenow fragment as a model for more complicated DNA polymerases.

The Figure (right) shows the structure of the Klenow fragment complexed with a fragment of DNA containing a mismatched terminal base pair. The 3′ end of the nascent strand is positioned at the 3′ → 5′ exonuclease site of the enzyme. During polymerization, the template strand occupies the groove at the top of the structure and at least 10 bp of double-stranded DNA are bound by the enzyme, as shown in the figure. Many of the amino acid residues involved in binding DNA are similar in all DNA polymerases, although the enzymes may be otherwise quite different in three-dimensional structure and amino acid sequence.

The unique 5′ → 3′ exonuclease activity of DNA polymerase I removes the RNA primer at the beginning of each Okazaki fragment. (Since it is not part of the Klenow fragment, the 5′ → 3′ exonuclease is not shown in the Figure above, but it would be located at the top of the structure, next to the groove that accommodates the template strand.) As the primer is removed, the polymerase synthesizes DNA to fill in the region between Okazaki fragments, a process called nick translation (see Figure below). In nick translation, DNA polymerase I recognizes and binds to the DNA chain. In this way, the enzyme moves the nick along the lagging strand. After completing 10 or 12 cycles of hydrolysis and polymerization, DNA polymerase I dissociates from the DNA, leaving behind two Okazaki fragments that are separated by a nick in the phosphodiester backbone. The removal of RNA primers by
DNA polymerase I is an essential part of DNA replication because the final product must consist entirely of double-stranded DNA.





The last step in the synthesis of the lagging strand of DNA is the formation of a phosphodiester linkage between the 3′-hydroxyl group at the end of one Okazaki fragment and the 5′-phosphate group of an adjacent Okazaki fragment. This step is catalyzed by DNA ligase. The DNA ligases in eukaryotic cells and in bacteriophage-infected cells require ATP as a cosubstrate. In contrast, E. coli DNA ligase uses NAD⁺ as a cosubstrate. NAD⁺ is the source of the nucleotidyl group that is transferred, first to the enzyme and then to the DNA, to create an ADP-DNA intermediate.

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[©Laurence A. Moran and Pearson/Prentice Hall]

Horton, H.R., Moran, L.A., Scrimgeour, K.G., Perry, M.D. and Rawn, J.D. (2006) Principles of Biochemistry. Pearson/Prentice Hall, Upper Saddle River, NJ (USA)