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Wecb 378
Wecb 378
Ronald Breslow,
New York
doi: 10.1002/9780470048672.wecb378
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Natural Products
Chemists have learned to imitate the principles of biologic chemistry in the quest to produce articial enzymes, articial receptors, and, ultimately, articial living cells. The eld of natural products chemistry is concerned chiey with discovering the structures of the molecules produced in living systems as well as the paths by which they are produced. Biomimetic chemistry takes nature as a model and extends what exists in biology into what is possible. This process includes using hydrophobic effects in water to achieve selective reactions and making new amino acids and new nucleotides to generalize the properties of proteins and nucleic acids. Potent new catalysts are based on the chemistry performed by coenzymes using bifunctional acidbase catalysis and the water exclusion achieved by natural enzymes. Exciting new areas include molecular machines, mimics of the chemical senses, and work on the origin on earth of the homochirality observed in biomolecules. The studies using nature as a model have enriched chemistry greatly and furnished interesting insights into biochemistry.
Since the beginning of time, people have learned about the world by observing nature and by imitating aspects of what we have learned. This observation is true particularly of what we see in living creatures. For example, people observed birds and insects in ight and dreamed of being able to y. Our most important clue was wings . We adopted this idea in the successful invention of airplanes. However, birds and insects power their ight by apping the wings; this concept proved less useful. Nature has limitations, and one of them is that a powered propellerand even more so a jet enginewere not practical for living creatures. Thus, we did not mimic birds slavishly; we adopted the central idea of their ability to y and added ways to power the ight that nature could not achieve. As Philip Ball said (1): A jumbo jet is not just a scaled up pigeon. Chemists have also taken many lessons and inspirations from natural chemistry; but, again, they have adapted their ideas so as not to be limited by the special requirements of natural chemistry. For example, nature designs proteins in three dimensions by folding linear polypeptide chains. This design indicates that genetic information is one dimensional, as is the sequence of nucleotides in genes. Although it might be interesting to use such a scheme in a mimic of an enzyme, chemists generally plan and then synthesize their three-dimensional structures, an option not open to nature.
Natural Products
In this brief article, I will focus on the ways we have learned from the chemistry of the life process itself. However, I must rst call attention to another part of natural chemistry that has an effect on chemical thinking: the eld called the Chemistry of Natural Products. Chemists have explored nature and have discovered several special types of chemicals: sugars, fats, terpenes, alkaloids, acetogenins, amino acids, heterocyclic coloring matters, macrocyclic lactones, steroid hormones, and so on. (2). These compounds have been produced by living organisms and they are secreted by or isolated from them. This eld is concerned with the substances of nature and the biochemical processes by which they are formed. Chemists have devoted much effort to exploring this natural world of chemistry as well as to determining structures; the natural world has stimulated the extension of the chemical world into models and analogs of the natural chemicals. The eld of organic chemistry was inuenced heavily by the types of chemical structures found in natural products; many medicinal compounds are still invented by using natural products as models for analogs. Chemists have also invented important polymers once nature showed us the natural polymeric carbohydrates, polypeptides, nucleic acids, and the polymers such as rubber that are produced from natural materials. 1
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Biomimetic chemistry
The eld concerned directly with imitating the chemistry of life processes has been called biomimetic chemistry or bio-inspired chemistry. The two terms differ in how close the mimic is to the natural process. As the author of this article, I will favor biomimetic, a word I coined (3).
Chemistry in water
A most important difference between human-made chemistry and living chemistry is that the latter occurs in water. Most chemical processes use organic solvents, which dissolve the components better; but water has special properties that are appreciated increasingly. Most signicantly, the hydrophobic effect in water is the force that causes membranes, micelles, and liposomes to form, which pushes hydrocarbon chains together to diminish the high-energy hydrocarbon/water interface. The hydrophobic effect is also a major reason that hormones bind to receptors and substrates bind to enzymes, and it causes the folding of polypeptide chains into the three-dimensional structure of proteins by promoting the clustering of hydrocarbon side chains in the protein interior away from water. Chemists now use water as a solvent for synthetic reactions, which takes advantage of the selectivities that the hydrophobic effect can induce (4). Many enzyme mimics (vide infra ) also use water as the medium to promote substrate binding into the catalyst. Of course, water as a solvent also has important environmental advantages over volatile organic solvents. It remains to be seen how much impact the special properties of water as a solvent will have within chemical synthesis and manufacturing.
well other related structures could perform the function of their natural models. In other words, why were these particular biologic molecules selected? A good example is the study of generalized forms of DNA. In one example, some of the heterocyclic bases were modied drastically and were replaced by substituted benzene rings, and yet the base-pairing process of DNA was still observed (6). In other studies, the deoxyribose of DNA was replaced by other sugars, in some cases with an improvement in binding properties (7). As such, the 2-deoxyribose in DNA was replaced by the isomeric 3-deoxyribose to observe how the binding properties were affected (8). Interestingly, the resulting isoDNAs did not pair well with their isoDNA partners, or with the normal DNA partner that had the appropriate base sequences. This result claried why 2-deoxyribose was selected for DNA even though the 3-deoxy isomer could be formed more easily under prebiotic conditions. However, the isoDNA base did pair well with its conjugate RNA, which furnishes insight into the difference between DNADNA pairing and DNARNA pairing (9). The coenzyme thiamine pyrophosphate (1) plays a central role in many parts of metabolism (Fig. 1). Its mechanism of action involves the formation of a thiazolium zwitterion 2 that was stabilized by a carbene resonance form 3 (10). This discovery opened up studies of the chemistry that such stabilized carbenes could catalyze, as chemists realized that the otherwise impossible chemistry that thiamine pyrophosphate catalyzes in nature could be generalized and adapted for useful synthetic processes. However, another study was an example of nature appreciationthe structure of thiamine was varied to learn what was special about the particular thiazolium derivative that was natural thiamine (11). As a chemical catalystignoring the question of what effect changes would have on the ability of the coenzyme to bind to the proteins that have evolved to use itthiamine proved to be the optimal relative to other related structures because of a balance of catalytic ability and chemical stability. The anion 4 derived from an imidazolium ring instead of a thiazolium ring was a weaker catalyst but was more stable in water (10). [The imidazolium anion and its dihydro derivative have proven to be very useful metal ion ligands, including
Nature appreciation
Another kind of generalization of natural chemistry involves viewing the natural chemistry in context by observing how 2
Figure 1 Thiamine, vitamin B1, is the cofactor, as its pyrophosphate ester, for many important biologic reactions. These reactions involve the formation of an anion 2 that is stabilized by resonance with a carbene form 3. The related species 4 and derivatives have been developed as important ligands for metal ions in chemical synthesis.
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in catalysts for olen metathesis (12). This chemistry was developed as an outgrowth of the understanding of how nature uses thiamine in biology.]
as two enantiomers. Again, the secret in enzymatic reactions is geometric control of the processes where the chiral centers are produced because of the geometry of the enzymesubstrate complex. Such enantiomeric selectivity is the target of much modern synthesis. A particular example from our laboratory is observed in compound 5, in which a basic amino group held rigidly on a mimic of the coenzyme pyridoxamine phosphate is able to convert ketoacids to amino acids with high enantioselectivity (15) (Fig. 3). This reaction is modeled closely to the way in which transaminase enzymes achieve the same goal.
Figure 2 In the rst steps of the conversion of lanosterol to cholesterol, an enzyme, cytochrome P-450, oxidizes the three methyl groups that will be removed while leaving the rest of the molecule untouched, because of geometric control that is typical in enzymes. WILEY ENCYCLOPEDIA OF CHEMICAL BIOLOGY 2008, John Wiley & Sons, Inc.
Figure 3 A mimic of the enzyme transaminase achieves high stereoselectivity in the product amino acid because of geometric control by the attached basic chain.
Figure 4 The enzyme ribonuclease A cleaves RNA using the imidazole group of histidine 12 as a base and the imidazolium group of histidine 119 as an acid in a simultaneous two-proton transfer process. Studies discussed in this article indicate that the process is not as simple as shown.
to the phosphorus, and the other one, protonated, which functioned as a proton donor group in the hydrolysis reaction. The best catalyst in this process was one in which the proton was being delivered to the phosphate anion group, which formed a ve-coordinate phosphorane intermediate. In a subsequent fast step, this went to the nal hydrolysis product, with the proton ending up on the OH group that is formed in the hydrolysis. We also examined the reaction rate in different mixtures of D2 O and H2 O, and we saw that the proton transfers were simultaneous, not sequential (19). Remarkably, the detailed data were almost identical with those observed in the enzyme process, which raised the question of whether the enzyme also used a mechanism proceeding through a ve-coordinate phosphorane intermediate, rather than going to the ring-opened product directly. We examined another biomimetic system to explore this question. We studied the reaction of a tiny fragment of RNA, uridyluridine 6, catalyzed by high concentrations of imidazole buffer (20) (Fig. 5). This buffer imitated the imidazole and imidazolium groups of the enzyme. In the enzyme, the effective concentration of these groups is high because of substrate binding right next to them. The products of the buffer-catalyzed reaction were uridine 7 and uridine cyclic phosphate 8, just as in an enzymatic cleavage process. However, we also saw that the buffer catalyzed an isomerization of the phosphate diester 6originally attached to the 3-oxygen of uridineto an isomer 9 in which it had migrated to the 2-oxygen. The chemistry of phosphate reactions, and in particular the nonequivalence of the ve groups in a phosphorane, requires 4
that such a migration proceed through a phosphorane intermediate that undergoes pseudo-rotation to permit the migration (21). Our other kinetic studies made it clear that the cyclization process went through the same phosphorane intermediate. We have proposed that the normal enzymatic hydrolysis process also proceeds through such a phosphorane intermediate, but isomerization does not occur because pseudo-rotation is not permitted in the enzyme (22). Complete agreement does not exist with our proposal (23). In this case, the biomimetic model system was not based only on natures enzyme, it pointed to possible aspects of the natural enzyme that had not been considered previously. Thus we had learned something from nature, and we returned the favor by learning something that could give insight into nature itself.
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Figure 5 Imidazole buffer catalyzes both the hydrolysis of uridyl-uridine, a simple RNA fragment, and also its isomerization. This nding indicates that a phosphorane intermediate is involved, and it suggests that such a process may also be used by the enzyme ribonuclease itself.
Generally, their studies involved hydrolytic and fragmentation reactions. We took up the study of such systems as mimics for transaminase enzymes and showed with the polyaziridines that we achieve very large accelerations of the conversion of ketoacids to amino acids (27). This result reected several ways in which these polyamines mimic enzymes. First of all, transaminations involve many steps in which protons are added and subtracted from the reacting species and the reaction intermediates. In enzymes such as ribonuclease, normally imidazoles are used to perform such proton transfers. Their pKas are close to the operating pH for the enzyme, which makes them both the strongest base and the strongest acid that can exist in free form. A stronger base would be protonated by the medium and a stronger acid would be deprotonated. It can be shown that the catalytic result of increased acid or base strength does not make up for the loss of concentration of the free acid and base species. The polyaziridines have their nitrogens so close to each other that they titrate over the range of pH 13 to pH 3, so they are half-protonated at pH 8 and have the strongest base and acid groups possible, just as in enzymes that use imidazole with a pKa near neutrality. Of course the polyaziridines achieve this not with special basic groups, just with the effect of neighboring charges that make it harder to add more positive charge. We also saw that the potency of the polyaziridines as enzyme mimics was increased strongly when hydrophobic chains were added to some of the nitrogens (described in Refs. 2427 and 2830), which was evident in two ways. First of all, with the hydrophobic core that these chains produced, we achieved strong binding in water solution of ketoacids that carried hydrophobic components. The binding was much better, as reected in a Michaelis Km , for indolepyruvic acidthat produced tryptophan on transaminationthan for pyruvic acid that produced alanine, which is not surprising. Perhaps more interestingly, the hydrophobic chains increased the rate constants signicantly for reaction of the complex (28), which reects a medium effect that is invoked often for natural enzymes. Water is to some extent an enemy of rate because acids, bases, and substrates often need to lose their bound waters to react. In nature, this problem is solved by performing the reactions in the hydrophobic interior of the enzymes away from the external water. The hydrophobic chains in our enzyme mimic performed the same function, which excluded water from the reaction site.
The transformations themselves involved reactions of ketoacids with a pyridoxamine unit, either covalently attached to the polymer or reversibly bound to the hydrophobic core (29), which converted the ketoacids to amino acids, and the pyridoxamine was converted to a pyridoxal unit either covalently attached to the polymer or reversibly dissociated from the polymer. This reaction was modeled directly on the transamination process observed in natural enzymes. However, the second part of a full transamination in nature is the reaction of the pyridoxal with a different amino acid, which runs the transamination backward to form the pyridoxamine again while converting the new amino acid into its corresponding ketoacid. We found that such a process was too slow in our biomimetic system and could not compete with the rapid aldol condensation of the ketoacids with the pyridoxal. To solve this problem, we used a mimic of a different enzyme, dialkylglycine decarboxylase (30). In this enzyme, pyridoxal phosphate reacts with an alpha-disubstituted glycine to perform an irreversible decarboxylation (Fig. 6) while converting the pyridoxal species to a pyridoxamine. We imitated this with our model transaminations using pyridoxal species that carry hydrophobic chains, and we were able to achieve as many as 100 catalytic turnovers. Thus, we could imitate one enzymethe ordinary transaminasesby also imitating another enzyme that solved the turnover problem. As we read more about disubstituted glycines, we learned that some such species are delivered by carbonaceous chondritic meteorites, and with partial enantioexcesses. This information offered a clue as to how biologic homochirality could have originated on earth (see below).
Figure 6 The mechanism used in the oxidative decarboxylation of alpha disubstituted glycines by an enzyme, which, in mimics, solved the problem of converting pyridoxal species to pyridoxamine species in biomimetic transaminase systems.
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Molecular machines
Living systems can convert chemical energy into mechanical motion. The most obvious examples are the motion in muscle contraction and the motion exhibited by agella in bacteria and in sperm cells. These models from nature have stimulated a few chemists to try to mimic such properties with promising results (31, 32). We do not yet know what practical benets such mimics will bring, but the creation of molecular machines by imitating those in nature is an exciting adventure. It is part of the fundamental change that chemistry itself is undergoing. Instead of being concerned only with the properties of pure chemical substances, the chemistry of the future will be concerned with the properties of organized multimolecular systems. It is an exciting prospect, which is stimulated by our observation of the wonderful properties of natures organized multimolecular system: the living cell.
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as a 95 to 5 ratio of L-phenylalanine starting with only a 50.5 to 49.5 ratiounder credible prebiotic conditions (38, 39). Such studiesand others attempting to understand how spontaneous chemistry in the primitive earth could have formed living cellsare not inspired directly by information about how life originated, but they are inspired by nature as we see it, which is populated as it is by life forms. The work does not aim to show how life originated, which is an historical question. However, it tries to develop the scientic evidence that can support a reasonable hypothesis of how it may have started.
Articial life
Another important goal of modern chemistry is directly inspired by nature: the hope to produce articial life, or at least some systems that share many of the attributes of life. The ultimate goal of biomimetic chemistry is to mimic life itself, not in the form that it now has but in alternate forms. Chemists have generalized so many types of substancesnovel polymers, novel carbohydrates and amino acids, novel hormones and enzyme inhibitorsthat to generalize natural chemistry is one of their most characteristic activities. Can they generalize life itself? Time will tell.
Other authors
A good fraction of modern chemistry, which includes medicinal chemistry, derives some of its inspiration from a consideration of nature as a model. It is impossible, in this brief article, to describe more than a fraction of the efforts. For additional work in the spirit of biomimetic chemistry, the following is a list of some authors not yet mentioned whose independent work should be consulted. Even here, this list is certainly not complete: Jacqueline Barton, Steven Benkovic, Albrecht Berkessel, Thomas Bruice, Jik Chin, Jean Chmielewski, E. J. Corey, Donald Cram, Peter Dervan, Francois Diederich, Mark Distefano, Jean Frechet, Samuel Gellman, John Groves, Andrew Hamilton, Donald Hilvert, Barbara Imperiali, Makoto Komiyama, Jean-Marie Lehn, Arthur Martell, Julius Rebek, Jean-Pierre Sauvage, Alanna Schepartz, Dieter Seebach, Donald Tomalia, and Steven Zimmerman.
References
1. 2. 3. 4. 5. Ball P. Stories of the Invisible: A Guided Tour of Molecules. 2001. Oxford University Press, Oxford, United Kingdom. Nakanishi K, Goto T, Ito S, Natori S, Shigeo N, eds. Natural Products Chemistry. 1974. Academic Press, New York. Breslow R. Biomimetic chemistry, centenary lecture. Chem. Soc. Rev. 1972;1:553580. Breslow R. Hydrophobic effects on simple organic reactions in water. Acc. Chem. Res. 1991;24:159164. Liu W, Brock A, Chen S, Chen S, Schultz PG. Genetic incorporation of unnatural amino acids into proteins in mammalian cells. Nature Meth. 2007;4:239244. Krueger AT, Lu H, Lee AHF, Kool ET. Synthesis and properties of size-expanded DNAs: toward designed, functional genetic systems. Acc. Chem. Res. 2007;40:141150. Eschenmoser A. Searching for nucleic acid alternatives. Chimia 2005;59:836850.
6.
8. Dougherty JP, Rizzo CJ, Breslow R. Oligodeoxynucleotides that contain 2 ,5 linkages: synthesis and hybridization properties. J. Am. Chem. Soc. 1993;114:62546255. 9. Sheppard TL, Breslow RC. Selective binding of RNA, but not DNA, by complementary 2 ,5 -linked DNA. J. Am. Chem. Soc. 1996;118:98109811. 10. Breslow R. On the mechanism of thiamine action. IV. Evidence from studies on model systems. J. Am. Chem. Soc. 1958;80:37193726. 11. Breslow R, McNelis E. Studies on model systems for thiamine action. Synthesis of reactive intermediates, and evidence on the function of the pyrimidine ring. J. Am. Chem. Soc. 1959;81:30803082. 12. Vougioukalakis GC, Grubbs RH. Ruthenium olen metathesis catalysts bearing an N-uorophenyl-N-mesityl-substituted unsymmetrical N-heterocyclic carbene. Organometallics 2007;26: 24692472. 13. Breslow R, Yang J, Yan J. Biomimetic hydroxylation of saturated carbons with articial cytochrome P-450 enzymesliberating chemistry from the tyranny of functional groups. Tetrahedron 2002;58:653659. 14. Yang J, Gabriele B, Belvedere S, Huang Y, Breslow R. Catalytic oxidations of steroid substrates by articial cytochrome P-450 enzymes. J. Org. Chem. 2002;67:50575067. 15. Zimmerman SC, Czarnik AW, Breslow R. Intramolecular general base-acid catalysis in transaminations catalyzed by pyridoxamine enzyme analogues. J. Am. Chem. Soc. 1983;105:16941695. 16. Richards FM, Wycoff HW. Bovine Pancreatic Ribonuclease. The Enzymes, Volume 4. 3rd edition. 1971. Academic Press, New York. pp. 647806. 17. Matta MS, Vo DT. Proton inventory of the second step of ribonuclease catalysis. J. Am. Chem. Soc. 1982;87:53165318. 18. Anslyn E, Breslow R. Geometric evidence on the ribonuclease model mechanism. J. Am. Chem. Soc. 1989;111:59725973. 19. Anslyn E, Breslow R. Proton inventory of a bifunctional ribonuclease model. J. Am. Chem. Soc. 1989;111:89318932. 20. Breslow R, Xu R. Quantitative evidence for the mechanism of RNA cleavage by enzyme mimics. Cleavage and isomerization of UpU by morpholine buffers. J. Am. Chem. Soc. 1993;115:1070510713. 21. Breslow R. Kinetics and mechanism in RNA cleavage. Proc. Nat. Acad. Sci. U.S.A. 1993;90:12081211. 22. Breslow R, Dong SD, Webb Y, Xu R. Further studies on the buffer-catalyzed cleavage and isomerization of uridyluridine. Medium and ionic strength effects on catalysis by morpholine, imidazole, and acetate buffers help clarify the mechanisms involved and their relationship to the mechanism used by the enzyme ribonuclease and by a ribonuclease mimic. J. Am. Chem. Soc. 1996;118:65886600. 23. Herschlag D. Ribonuclease revisited. Catalysis via the classical general acid-base mechanism or a triester-like mechanism. J. Am. Chem. Soc. 1994;116:1163111635. 24. Klotz IM, Suh J. Evolution of synthetic polymers with enzyme-like catalytic activities. In: Articial Enzymes. Breslow R, ed. 2005. Wiley-VCH. Weinheim, Germany, pp. 6388. 25. Suh J. Synthesis of polymeric enzyme-like catalysts. Synlett 2001;9:13431363. 26. Hollfelder F, Kirby AJ, Tawk DS. Efcient catalysis of proton transfer by synzymes. J. Am. Chem. Soc. 1997;119:95789579. 27. Liu L, Breslow R. Vitamin B6 enzyme models. In: Articial Enzymes. Breslow R, ed. 2005. Wiley-VCH. Weinheim, Germany, pp. 3762.
7.
WILEY ENCYCLOPEDIA OF CHEMICAL BIOLOGY 2008, John Wiley & Sons, Inc.
28.
29.
30.
31.
32.
33.
34. 35.
36. 37.
38.
39.
Liu L, Rozenman M, Breslow R. Hydrophobic effects on rates and substrate selectivities in polymeric transaminase mimics. J. Am. Chem. Soc. 2002;124:1266012661. Liu L, Zhou W, Chruma JJ, Breslow R. Transamination reactions with multiple turnovers catalyzed by hydrophobic pyridoxamine cofactors in the presence of polyethylenimine polymers. J. Am. Chem. Soc. 2004;126:81368137. Chruma JJ, Liu L, Zhou W, Breslow R. Hydrophobic and electronic factors in the design of dialkylglycine decarboxylase mimics. Bioorg. Med. Chem. 2005;13:58735883. Kelly TR, Cai X, Damkaci F, Panicker SB, Tu B, Bushell SM, Cornella I, Piggott MJ, Salives R, Cavero M, Zhao Y, Jasmin S. Progress toward a rationally designed, chemically powered rotary molecular motor. J. Am. Chem. Soc. 2007;129:376386. Pijper D, Feringa BL. Molecular transmission: controlling the twist sense of a helical polymer with a single light-driven molecular motor. Angew. Chem. Int. Ed. 2007;46:36933696. Axel R. Scents and sensibility: a molecular logic of olfactory perception (Nobel lecture). Angew. Chem. Int. Ed. 2005;44: 61116127. Buck LB. Unraveling the sense of smell (Nobel lecture). Angew. Chem. Int. Ed. 2005;44:61286140. Anslyn EV. A marriage of supramolecular chemistry with pattern recognition. 232nd ACS National Meeting, San Francisco, CA, 2006. Pizzarello S. The chemistry of lifes origin: a carbonaceous meteorite perspective. Acc. Chem. Res. 2006;39:231237. Breslow R, Levine M. Partial transfer of enantioselective chiralities from alpha-methylated amino acids, known to be of meteoritic origin, into normal amino acids. Tetrahedron Lett. 2006;48:18091812. Breslow R, Levine M. Amplication of enantiomeric concentrations under credible prebiotic conditions. Proc. Nat. Acad. Sci. U.S.A. 2006;103:1297912980. Klussman M, Iwamura H, Matthew S, Wells D, Pandya U, Armstrong A, Blackmond D. Thermodynamic control of asymmetric amplication in amino acid catalysis. Nature 2006;441:621623.
See Also
DNA-Binding Molecular Motors Enzyme Cofactors, Chemistry of Life, Origins of Organic Chemistry in Biology Water, Properties of
Further Reading
Breslow R. Studies in biomimetic chemistry. Pure & Appl. Chem. 1998;70:267270. Breslow R. Imitating antibodies and enzymes. Proc. Robert A Welch Foundation 40th Conference on Chem. Res. 1997;40:111. Breslow R. Biomimetic chemistry and articial enzymes: catalysis by design. Accts. Chem. Res. 1995;28:146153. Breslow R. Binding and catalysis in water. Supramol. Chem. 1992; 411428. Breslow R. Hydrophobic effects on simple organic reactions in water. Accts. Chem. Res. 1991;24:159164.
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