Phosphorus Chemistry - ACS Publications - American Chemical Society

Chapter 1 The Role of Phosphorus in Chemistry and Biochemistry An Overview F. H. Westheimer Downloaded by COLUMBIA UNIV on July 23, 2013 | http://pubs.acs.org Publication Date: April 7, 1992 | doi: 10.1021/bk-1992-0486.ch001

Department of Chemistry, Harvard University, Cambridge, MA 02138

Ionized phosphate esters are ubiquitous in biochemistry principally for two reasons. First, metabolites generally must be charged, so that they will not pass through a lipid membrane and so be lost to the cell. Second, the charge must be negative, so as to repel nucleophiles and thus resist destruction by hydrolysis. Phosphates uniquely allow for these requirements. We cannot overestimate the importance of phosphorus to chemistry and biochemistry; its central place has been quietly acknowledged, especially by the work that molecular biologists carry out daily. Figure 1, courtesy of Professor George Kenyon, illustrates the position of phosphorus in the Periodic Table. INDUSTRIAL CHEMISTRY Phosphorus chemistry has proved a minor branch of traditional organic chemistry and of modern inorganic chemistry, but it is of great importance in industry, and especially in biochemistry and molecular biology. The greatest tonnage of industrial phosphorus chemicals is undoubtedly superphosphate, used as fertilizer; the phosphate is incorporated into growing plants. The most ubiquitous phosphorus product of industry is certainly matches, where phosphorus sesquisulfide (P4S3) as fuel, along with potassium chlorate as oxidizer, provides the incendiary mixture; this mixture loses none of its importance because the combination is a very old discovery. Phosphorus compounds are also important in foodstuffs — for example, phosphoric acid acidifies soft drinks — and in plasticizers, and in fire retardants; phosphorus compounds are used as war gases, a fact that many of us feared would become all too evident in the Gulf War. Arthur Toy's books, entitled Inorganic Phosphorus Chemistry and Phosphorus Compounds in Everyday Living, have been a principal source of information about industrial phosphorus chemicals for countless students. The 0097-6156/92/0486-0001$06.00/0 © 1992 American Chemical Society In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.

Downloaded by COLUMBIA UNIV on July 23, 2013 | http://pubs.acs.org Publication Date: April 7, 1992 | doi: 10.1021/bk-1992-0486.ch001

2

PHOSPHORUS CHEMISTRY

H ϋ

ft*

Β

No

*fl

Al

Κ Co Se Τι

V Cr Mn Fe Co

Kb Sc Y

Zr

Nb Mo Tc Ru Rh

Ct i o Ι ο

Hf

Τα W Re

Ot

Ir

C

Si

Ni Cu Zn G a G e Pd Ag cd In Pt Au

Sn

τι P b

Ν

P As

o s

H He F Ht Cl Αι

Se Br le I Po

Fr Ko Ac

•fc.

Fig. 1. The position of phosphorus in the Periodic Table.

In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.

At

Kr K« Μη

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WESTHEIMER

The Role of Phosphorus in Chemistry and Biochemistry


symposium in his honor allows us to acknowledge the role he has played over the years, especially in popularizing the importance of phosphorus chemistry. BIOLOGICAL CHEMISTRY M y personal interest in phosphorus chemistry is largely directed toward its role in biochemistry. I first became acquainted with the importance of the field of nucleotides because of a series of lectures that Lord Todd gave at the University of Chicago in 1948, and I am forever grateful to him for the exciting and inciteful introduction to the field that he provided. The role of phosphates in biochemistry was foreshadowed by the use of phosphates as fertilizer; the occurrance of phosphates in plants implied an important role of phosphates — or anyway of phosphorus — in the processes of life.H) D N A . The most spectacular role of phosphates in life processes is as the central building block in nucleic acids. The non-scientific public — that is to say, almost everybody — is well acquainted with the concept that the genetic material is D N A , or deoxyribonucleic acid, but relatively few non-scientists know that D N A is a chain of diesters of phosphoric acid. Philip Handler told the story of a young man whom he met on an airplane, who told him that he knew all about nucleic acids, and really had only one question: What is an acid? But phosphates are also involved in ATP, that is in adenosine triphosphate, and in other compounds for the storage of chemical energy. Further, phosphate residues are attached to several coenzymes, and phosphate residues, combined with the hydroxyl groups of the serine and threonine and tyrosine residues of enzymes, control the action of these catalysts.(2) A n understanding of the fundamental chemistry of phosphorus allows us to understand the chemistry of these biologically important materials. In 1987,1 offered a view of why Nature chose phosphates for the genetic tape, and for the storage of chemical energy, and for control processes, and so on. THE IMPORTANCE OF BEING IONIZED The most important aspect of my analysis was derived from a paper that Bernard David (3) published in 1958, entitled "The Importance of Being Ionized." Cells — presumably including the earliest cells on earth — are distinguished by lipid membranes. Davis pointed out that ionized compounds generally cannot pass through lipid membranes, whereas most electrically neutral compounds do so; the highly polar ions do not dissolve in the nonpolar fatty acid residues of the membrane. Therefore, most metabolites, if they are to be retained within cells, must be ionized. Of course, there are some compounds — a minority of them — that can get along without being ionized. Steroids and other compounds that are almost totally insoluble in water may dissolve in the membrane but will not pass through to a surrounding watery environment. (Figure 2)


3


4


Fig. 2. The escape of unionized molecules from a cell membrane Some sugars and other compounds with many hydroxyl groups may be so insoluble in the membranes that, once they are generated within a cell, they will remain inside, and will not escape to the outside, watery environment. But many, and perhaps most unionized compounds will dissolve to a sufficient extent in a membrane to pass through it, and be lost to the cell. Primitive organisms depended for survival on keeping important metabolites within the cell membrane, and an important way to do this, perhaps the only practical way to do this for many compounds, was to manage somehow to convert the metabolite into an ionized compound. Phosphoric acid is a moderately strong acid; it and its mono- and diesters show an ionizable proton with a pK of about 2. It follows that mono- and diesters of phosphoric acid will be almost completely ionized in aqueous biological media where the p H is usually somewhere near neutrality. They will, therefore, be retained within a cell membrane. A n effective way to convert a metabolite to an ionized compound, and thus retain it within a cell membrane, is to attach a phosphate residue to it. This is precisely what has happened, what has evolved. A large number of metabolites, including several coenzymes, are phosphate esters. In particular, the genetic tape consists of phosphate diesters. The tape that has evolved is built of units — the nucleoside phosphates — that can be assembled and then disassembled, and reused. This seems an efficient plan; the nucleosides are complex structures that require considerable metabolic energy to build. If one is to use nucleosides at all, they ought not be wasted. This argument assumes that the genetic tape will be constructed of nucleosides. The argument, and the



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interests of this symposium, are concerned, among other things, with the question of why Nature chose phosphates, but not why it chose nucleosides. That is an interesting question, too, but separate from the question of phosphates, and deserves a symposium of its own. If one makes the genetic tape from nucleosides, one must link them together to make the tape. The best way to do this is to take advantage of the hydroxyl groups in deoxyribose — or in ribose to make R N A — and to esterify these hydroxyl groups. In order to link the groups together, one will need a divalent linker, and to make esters, one will then need an acid that is at least dibasic. But if in addition, each unit must carry a charge, so as to retain the material within the cell membrane, then the acid must be tribasic. At least, this is true if the charge must be negative. The reason why the charge must be negative, and why a positive charge positively won't do, is the next topic of discussion. For the present, we note that, if one needs a tribasic acid, then the first compound that comes to mind is phosphoric acid. In a fuller discussion, one can show that no other tribasic acid, or at least none that readily comes to mind, will do. S T A B I L I T Y The next question with respect to the choice of phosphates for the genetic tape concerns stability, and considerations of stability dictate that the charge on the tape must be negative. A l l metabolites must be sufficiently stable to survive long enough to carry out their biological function. Many compounds are formed and used in brief periods of time, but this is not, and cannot be so for the genetic material; it must be chemically stable for a time comparable to the length of life of the organism in question. This may be less that an hour for bacteria and viruses, but must be many years — even decades, for mammals such as man. Furthermore, the genetic tape has many millions of ester bonds, and although the cleavage of some of them may be tolerated, still the cleavage of only one bond could be fatal. The genetic tape must then be extraordinarily stable against hydrolysis. D N A meets this requirement, and not many other esters do. Furthermore, the reason for its stability lies in the multiple negative charges it carries. Esters of ordinary carboxylic acids are insufficiently stable to be considered for the genetic material. Even at neutral pH, the half-life of a typical carboxylic acid ester in water at room temperature is only about a month, and one ester bond in a million — enough to break the tape in a hundred places — will hydrolyze every couple of seconds. The rate will be greater — much greater — in alkaline or acid solutions, and although biochemical environments tend toward neutrality, many of them differ significantly from that ideal. Triesters of phosphoric acid are somewhat more stable than esters of carboxylic acids, but nowhere near stable enough to serve as a genetic tape. (Figure 3.)



6

CH -C* OCoH 25 3

(CH 0) P=0 3

3

n

1 bond in 10 cleaved in water at 25° and pH 7 in « 2 seconds


6

1 bond in 10 cleaved in water at 25° and pH 7 in « 1 minute 6

Fig. 3. Rates of hydrolysis of ethyl acetate and trimethyl phosphate. But fortunately the tape is composed of diester monoanions of phosphoric acid. The tape is multiply charged, and all of these charges are negative. The charges not only perform the function of retaining the genetic material within the cell membrane, but stabilize it against hydrolysis. Similarly, the negative charges in R N A — ribonucleic acid — stabilize it against hydrolysis. Since ribonucleic acid is inherently much less stable than deoxyribonucleic acid, the stabilizing effect of negative charge is even more important here than with DNA. Negative charges repel nucleophiles, such as hydroxide ion and water, and slow the rate of hydrolysis enormously. The negative charges make possible the use of the diester monoanions of phosphoric acid for the genetic tape. E L E C T R O S T A T I C S Quantitatively, how great is the effect of negative charge on the rate of hydrolysis? The effect of a single nearby negative charge on the rate of attack of a negatively charged nucleophile, such as hydroxide ion, is enormous; the rate factor for phosphates is of the order of 10^. In general, the rate factor depends upon the closeness of approach of the nucleophile to the charge; the electrostatic effect can be small, or it can be enormous, even compared to 10 . A n estimate of the effect of a negative charge on a negatively charged nucleophile can be obtained by considering the ratio of the first to the second ionization constants of polybasic acids. This ratio is controlled by the electrostatic effect of a negative charge on a proton. Of course, the electrostatic free energy produced by the interaction of a proton and a negative charge will be of opposite sign to that produced 5


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by the interaction of two negative charges, but the absolute magnitude should be about the same. (Figure 4.)

H+

v

OH

CP*

x

O-H

d

+

^


Fig. 4. The electrostatic interaction of a proton with a negative charge in the ionization of a dibasic carboxylic acid. If we are dealing with a polybasic acid where all the ionizations occur from protons attached to oxygen, then die first ionizable proton is removed against the incipient negative charge of the nascent oxygen anion. The second ionizable proton is removed against a similar force from the nascent anionic charge on the oxygen atom to which it is originally attached, plus an additional electrostatic force arising from the interaction between the proton and the negative charge left behind by the first ionization. If one assumes that the free energy of ionization of a proton from the oxygen atom to which it is attached is roughly the same for the first and for the second ionizations, then the difference in free energy for the two ionizations is simply the electrostatic free energy that arises from removing a proton in the field of the charge left over by the first ionization. The smaller the distance between the second proton and the residual negative charge, the greater the effect. Thus the ratio of the first to the second ionization constants for phosphoric acid is 10^, whereas that for citric acid is only 50. These numbers need to be corrected for small statistical factors, but the statistical factors don't affect the general conclusion at all, i.e. that the electrostatic effect depends inversely upon the distance between the ionizing groups (Table 1). TABLE 1 Ionization Constants

K

Κ

λ

Phosphoric

Citric

7.5 χ 1 0

7.1 χ 10

3

K

2

6.2 χ 10"

1.7 χ 10

8

3>

molar

2.2 χ 10"

6.4 χ 10"


13

6



A similar effect controls the rate of attack of hydroxide ion on an ester. The attack of hydroxide ion on an electrically neutral ester, such as trimethyl phosphate, involves many interactions. The attack of hydroxide ion on an ester-anion, such as dimethyl phosphate anion, presumably involves the same interactions, plus an additional electrostatic interaction between the hydroxide ion and the residual negative charge on the phosphate anion. (Figure 5.)

OCH

3

OCH3

Fig. 5. Attack of hydroxide ion on dimethyl phosphate anion Since the ratio of the first to the second ionization constants of phosphoric acid is 10 , we would expect — and find — that the ratio of the rate constants for attack by hydroxide ion on trimethyl phosphate to that for attack on dimethyl phosphate anion is also about ΐΦ. Uncharged nucleophiles, such as water, are also repelled by negative charges, although of course less strongly. Nevertheless, the rate of attack of water on a negatively charged substrate is much less than that on an electrically neutral one. Any nucleophile, almost by definition, attacks an electrophile with a free electron pair, and this electron pair is repelled by negative charge. A negative charge, therefore, protects an ester against nucleophilic attack. A positive charge, by contrast, will attract nucleophiles, and a positive charge near the site of reaction makes an ester more susceptible to hydrolysis, and thus less, rather than more stable against hydrolysis. A n example of this effect will be presented later in this article. These facts and principles help explain why phosphoric acid is ideal for its role in DNA. Let me insert a personal footnote here. These electrostatic effects were first discussed in the chemical literature by Niels Bjerrum (5) in 1923, and in 1938 John Kirkwood and I developed a crude model for such electrostatic systems, and worked out an approximate mathematical theory to put these effects on a quantitative base/6,7,8) 5



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Isn't it astonishing that work I did in 1938, before I knew anything about the role of phosphates in biochemistry, should be pertinent to my own interests more than fifty years later.? When I was a graduate student at Harvard in the early 1930's, I read a paper by Arthur Michael — the Arthur Michael who invented the Michael condensation reaction — one of the early heroes of American organic chemistry. He was, at that time, a Professor Emeritus of Chemistry at Harvard. To the best of my recollection, Michael wrote that "Fifty years ago, my coworkers and I . . . . " (I have not yet found this exact quote, but Michael (9) refers to work he did (10) fifty-four years earlier.) At the time, this phrase seemed almost hysterically funny; imagine someone maintaining his interest in chemistry for over half a century! A t present, I am a Professor Emeritus of Chemistry at Harvard, writing that, more than fifty years ago, Jack Kirkwood and I developed a quantitative theory of electrostatic effects. Perhaps some readers will find this at least mildly amusing. M y guess, however, is that most chemists maintain an active interest in chemistry as long as they live, and that my reaction, back in the 1930's, just tells you how young I was then. In any event, the genetic material is stable against hydrolysis because of multiple negative charges. The negative charges do double duty; they both stabilize the genetic tape against hydrolysis and keep it safe within the lipid cell membrane. ATP The principal compound for the storage of chemical energy is adenosine triphosphate; like the genetic tape, it is stabilized against hydrolysis by its negative charges. The free energy for the hydrolysis (11) of ATP is -7.3 kcal/mole, as compared to only -2.2 kcal/mole for the hydrolysis of an ordinary phosphate ester. This excess free energy can be used in accomplishing the phosphorylation of various metabolites, or in supplying the needed driving force for many other reactions of biochemistry; the hydrolysis of ATP can be coupled with other metabolic processes that otherwise would be energetically unfavorable. A phosphorylation process involves the transfer of a monomeric metaphosphate unit, PO3-, from ATP to the substrate in question. (Figures 6 and 7.) PO3- is a strong electrophile, and a strong phosphorylating agent. In research in my laboratory, my collaborators have shown that, in hydrophobic environments methyl metaphosphate w i l l even phosphorylate the ring in aromatic amines (12,13) Furthermore, we have shown that monomeric metaphosphate ion, presumably but not necessarily free, attacks carbonyl groups, and will convert ketones to the corresponding enol phosphates.(24,15) W.P. Jencks (16) has shown that monomeric metaphosphate is never free in aqueous solution; in this regard, it resembles the proton, which is an even stronger electrophile, and similarly is never free in aqueous solution. In acid catalyzed reactions, a proton is transferred from one molecule to


9



10

OH (4 negative charges!) Fig . 6 Adenosine triphosphate.

[ATP] " + H 0 -> [ADP] * + H P0 " 4

2

3

2

4

AF = -7.3 kcal/mole

HO^^Y^OPOaH--^ HO

HO^V^

O H

+ H

2

P

HO

AF = -2.2 kcal/mole Fig.7. Thermochemistry of hydrolysis of ATP


°

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another; in fact, protons are probably completely free only in the gas phase. Monomeric metaphosphate is nowhere near as electrophilic as protons; nevertheless, Professor Jencks has shown that, at least in aqueous solution, monomeric metaphosphate, like the proton, is transferred from one molecule to another without ever being free. This is not necessarily the case in less nucleophilic media, and we — the scientific community — will try our best to determine whether the environment of enzymes resembles aqueous solution or a much less polar medium. Such investigations are of real scientific interest, but must not obscure the important fact that the role of monomeric metaphosphate in biochemistry is as an electrophile. Like the proton, its electrophilicity is sharply diminished by water and, at least in aqueous solution, it is transferred from one molecule to another without becoming free. Unlike the proton, which is never free in any liquid medium, monomeric metaphosphate is probably free in apolar solution; like the proton, its role is that of an electrophile. In any event, water is the enemy of monomeric metaphosphate. ATP, which transfers or generates PO3", depending on the medium, is thermodynamically quite unstable i n aqueous solution, but nevertheless kinetically stable. In 1951, Fritz Lipmann (17) noted that ATP is stabilized against hydrolysis by its negative charges. The compound occupies a unique place in biochemistry because it is thermodynamically unstable yet kinetically nearly inert — a marvelous and apparently incompatible set of properties that is related to the polybasic nature of phosphoric acid, and the resulting negative charges on ATP. SYNTHESIS OF D N A The last section of this report concerns the Letsinger-Caruthers synthesis of D N A . It seems to me that an introductory paper should concentrate on the most important topics, even if my own contribution to the topic is pretty small. Biochemists today isolate and replicate the genes for many enzymes, and then express and isolate proteins related to those genes. They can work up a piece of nucleic acid from a tiny sample — a few billion molecules — to make milligrams of nucleic acid and eventually express many milligrams of enzyme. Biochemists and molecular biologists perform these miracles routinely, in months or sometimes even in weeks. Fortunately for the progress of science, some chemists have joined in — in the Chemistry Department at Harvard alone, Stuart Schreiber, Gregory Verdine, George Whitesides, and — until he became the Dean of the Faculty of Arts and Sciences — Jeremy Knowles — together with their students — are using these techniques, and of course many more chemists are doing the same sort of thing elsewhere. The techniques allow site directed mutagenesis of enzymes, among other research directions, and this allows a much more exacting test of the detailed chemical


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mechanism of enzyme action. In this connection let me mention the use of site-directed mutagenesis by my former graduate students, John Gerlt and George Kenyon (18,19,20) in determining the mechanism of action of mandelate racemase. There are, of course, many more examples. Essential to these exercises in molecular biology is the ability to synthesize pieces of nucleic acid of specified sequence and reasonable length, that is, to make polynucleotide sequences thirty to a hundred nucleotides long. These pieces of synthetic nucleic acid are needed, among other uses, to fish out specific genes for which only a bit of sequence is known, and to fish them out from a biochemical soup of almost unbelievable complexity. Chemists now have automatic machines into which you can type the needed sequence, turn on the machine and walk away — though in fact you had best not walk away — and get a fifty-step synthesis in a day. The product can be purified by chromatography, and the sequence can readily be checked by the techniques for which Frederick Sanger and Walter Gilbert received the Nobel prize in 1980.(22,22) Molecular biologists who use this material seem to take the synthesis for granted. If you need a shirt, you go to a clothing store and buy one i n your size. If you need a specific polynucleotide, you go to a chemist, and he synthesizes it for you. There can't be much to it; a machine does it. But really, the machine is a miracle of chemical inventiveness. How many molecular biologists even recognize the names of Robert Letsinger and Marvin Caruthers, their benfactors? One famous practitioner, who generally acknowledges the importance of chemistry, accepted without question that chemists synthesize things, just as carpenters make tables; apparently there's nothing to it. L E T S I N G E R - C A R U T H E R S SYNTHESIS Letsinger's original synthesis (23) condensed phosphites with the hydroxyl groups of deoxyribosides to build a growing chain that resembles the genetic material except that the last residue is a phosphite, instead of a phosphate. The phosphite is then gently oxidized to a phosphate. The earlier syntheses of polynucleotides, pioneered by Alex Todd (24) and by H . Gobind Khorana (25) , lighted the way to modern methods, but although these earlier syntheses were absolutely essential to the development of the field, they are much too time-consuming to be really practical. One can understand the advantage of the phosphite method when one realizes how much more reactive phosphites are than phosphates. Specifically, the rate of acid hydrolysis of an ester of phosphorous acid — that is to say, a phosphite — is about 10*2 times as great as that for the corresponding phosphate (26) - a thousand billion times as


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great. This advantage corresponds to the best that enzymes can manage; enzymes usually increase reaction rates by a factor of 10^ 1()12 Smart chemistry makes this synthesis the equivalent of an enzymatic one. The Letsinger-Caruthers synthesis is unrelated to the biochemical one except in the area of efficiency. Because phosphites are so reactive, they would be totally unsuitable for the genetic tape itself, but because they are so reactive, they are ideally suited as intermediates in the chemical synthesis of polynucleotides. One can accomplish the needed condensation reactions with phosphites under strictly anhydrous conditions with very weakly acidic catalysts, and then stabilize the product by a mild oxidation of the phosphite to the desired phosphate with iodine. The essential feature of the phosphites — the reason why the phosphites are so susceptible to acid hydrolysis or alcoholysis, is that the phosphorus atom of the phosphite is blessed with an unshared electron pair. A proton can then add to the unshared electron pair of a phosphite, or of a phosphoramidite. (Phosphoramidites are discussed below.) The positive charge that results from attaching a proton directly on phosphorus atom sensitizes the phosphorus to nucleophilic attack. This is the same sort of electrostatic effect that is discussed above in connection with the retarding effect of a negative charge on the hydrolysis of phosphates, but the effect of a positive charge increases the rate of nucleophilic attack, and the effect is of course enormously greater because the positive charge is directly on phosphorus, whereas the negative charge in a phosphate anion is on an oxygen atom, one bond length removed from the phosphorus atom. (Figure 8.)


#

OR I :P-OR OR

OR I H-P-OR I OR

+

ι +

Fig. 8. Protonation of phosphites and phosphoramidites.


13


14


With courage and an equation for electrostatic potential, one could have predicted the approximate magnitude of this electrostatic effect, but in fact no one did. Although Letsinger's original phosphite synthesis is an efficient process, it was still too difficult for general use, as it required dry ice temperatures and strictly anhydrous conditions. The method, however, was modified by Marvin Caruthers (27) who substituted monoester diamidites for phosphites. The reactions proceed much more smoothly with the amidites, which are, relatively, resistant to hydrolysis and to autoxidation. A beginning of the physical organic chemistry of the phosphites has been published (22), but to the best of my knowledge none has been published for the phosphoramidites. Perhaps this review will stimulate someone to take up this problem,just as Todd's lectures stimulated me to work in this area. In some detail, the Letsinger-Caruthers synthesis involves attaching a nucleotide to a solid support, such as silica gel, and preparing protected phosphite reagents for the four nucleotides. A 2,4dimethoxytrityl group, which can be easily removed when desired, is usually used for temporary protections of the 5'-hydroxyl group of the nucleoside. Methoxyl groups, or more frequently cyanoethyl groups, are used to protect the phosphorus-atom during the synthesis. One such protected reagent is then allowed to react with the 5'-hydroxyl group of the nucleotide tethered to a solid support. The resulting phosphate-phosphite compound is then oxidized with iodine to produce a protected dinucleotide. The dimethoxytrityl group that protects the 5'-end of the tethered dinucleotide can then be removed, by mild acid treatment, and the chain lengthened by repeating the process with the desired phosphoramidite. Finally, at the end of the synthesis, the protecting methoxyl groups, or whatever groups have been used to protect the phosphates, must be removed, protecting groups must be removed from the bases, and the completed molecule must be liberated from the solid support. The individual steps can be carried out with 99% yields, and long polynucleotides, with fifty or more units, can be prepared in this way. Some details of the chemical synthesis are not fully understood; in particular, the physical-organic chemistry is still incomplete or missing. But even without a full understanding of all the details, the synthetic procedure works, and works wonderfully. The chemical community can be proud of this contribution of chemistry to molecular biology. (Figure 9.)



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DMT = Dimethoxytrityl R = /soPropyl

Silica Fig. 9. The Letsinger - Caruthers synthesis of a protected form of nucleotides. Continued on next page In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.

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16

Fig. 9. Continued Phosphorus chemistry and biochemistry are enormous topics, and obviously this essay touches on only a few aspects of the subject; many other aspects are discussed in detail in this volume by others. This introduction to the symposium is built around the commanding importance of electrostatics in the chemistry and biochemistry of phosphates and phosphites, but that emphasis necessitated omitting a discussion of many other topics. These include, for example, pseudorotation in the hydrolysis of cyclic esters of phosphorus, a fascinating topic to which Edward Dennis and David Gorenstein and others in my laboratory have made significant contributions (28). The brilliant work of Usher, Richardson and Eckstein (29) which (if you w i l l pardon a pun) ushered in the determinations of the stereochemistry at phosphorus in the reactions of phosphate esters, has been omitted, as has the work of Jeremy Knowles (30) and his coworkers, who demonstrated how to use Ο , Ο * , and O , to 1 6

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The Role of Phosphorus in Chemistry and Biochemis

produce chirality at phosphorus. I have completely ignored the role of phosphorylation in the control of enzymic processes. But perhaps the work cited demonstrates the importance and conveys the excitement of phosphorus chemistry, and so provides an introduction to this symposium.


LITERATURE CITED

(1) J. W. Mellor Treatise on Inorganic Chemistry, VIII, 736 ff Longmans-Green:(1928). (2) L. Stryer Biochemistry (3rd Ed.) W. W. Freeman, N,Y. (1988). (3) F.H. Westheimer, SCIENCE 1987, 235 173 (4) B. Davis Archives Biochem. Biophys. 1958, 78 497. (5) N. Bjerrum Z. physik. Chem. 1923, 106 219. (6) J. G. Kirkwood; F. H. Westheimer, J. Chem Phys 1938, 6, 506 (7) F. H. Westheimer; J. G. Kirkwood ibid 1938, 513. (8) F. H. Westheimer; M. W. Shookhoff J. Am. Chem. Soc. 1940, 62 269. (9) A. Michael; J. RossJ.Am. Chem. Soc. 1933, 55 , 3684. (10) A. Michael Am. Chem. J. 1879, 1 , 312. (11) L. Stryer, op. cit.p317. (12) C. H. Clapp; A. Satterthwait; F. H. WestheimerJ.Am. Chem. Soc. 1975, 97, 6873. (13) A. Satterthwait; F. H. Westheimer ibid. 1978, 100, 3197; 102 4464 (1980). (14) A. Satterthwait; F. H. WestheimerJ.Am. Chem. Soc. 1981,103 1177. (15) K. C. Calvo; F. H. Westheimer ibid. 1983, 105 , 2827. (16) W. P. Jencks Acc. Chem. Res. 1980, 13, 161 (17) F. Lipmann in Phosphorus Metabolism, W. D. McElroy and H. B. Glass, Eds.; Johns Hopkins Press: Baltimore, 1951, Vol I,p521. (18) V. M. Powers et al. Biochemistry, 1991, 30 9255 (19) D. J. Neidhart et al. ibid 1991, 30 9264 (20) J. A. Landro et al. ibid 1991, 30 9274 (21) A. M. Maxam; W. Gilbert Proc. Natl. Acad. Sci. 1977,74, 560 (22) F. Sanger; S. Nicklen; A. R. Coulson ibid , 5463 (1977). (23) R. L. Letsinger; W.B. LunsfordJ.Am. Chem. Soc. 1976, 98 3655 (24) A. Todd Prospectives in Organic Chemistry, (A. Todd, Ed.), Interscience, N.Y. 1956,p245. (25) H. G. Khorana Pure Appl. Chem, 1968,17, 349 (1968). (26) F. H. Westheimer; S. Huang; F. CovitzJ.Am. Chem. Soc. 1988, 110,181. Errata, Ibid, 2993. (27) M. H. Caruthers SCIENCE, 1985, 230 281. (28) F. H. Westheimer Acc. Chem. Res. 1968, 1 70. (29) D. A. Usher; D. I. Richardson, Jr.; F. Eckstein, NATURE (London), 1970, 228 663 (30) S. J. Abbott et al.,J.Am. Chem. Soc. 1978, 100 2558. RECEIVED December 17, 1991 In Phosphorus Chemistry; Walsh, E., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.

Phosphorus Chemistry - ACS Publications - American Chemical Society

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