BRUCE MERRIFIELD 1921-2006

A Biographical Memoir by

ULF RAGNARSSON

© 2013 National Academy of Sciences Any opinions expressed in this memoir are those of the author and do not necessarily reflect the views of the National Academy of Sciences. BRUCE MERRIFIELD Photo by the Nobel Foundation, courtesy AIP Emilio Segre Visual Archives. July 15, 1921–May 14, 2006

BY ULF RAGNARSSON

BRUCE MERRIFIELD, John D. Rockefeller Jr. Professor Emeritus at Rockefeller 1 University and winner of the 1984 in Chemistry, was a prominent scientist with major influence on the development of peptide and protein synthesis, a field of great importance during the second part of the twentieth century. His name is associated with a fundamentally novel scientific method that he conceived, pioneered, and, during the rest of his life, further improved. Under his direction, this method was applied to problems of immense complexity. Bruce was born in Fort Worth, Texas, the only son of George Evan Merri- field, an interior decorator and furniture salesman, and Lorene Lucas Merrifield. The family went to Southern California in 1923 and moved a number of times during the Great Depression. As a result, Bruce attended eleven schools before bruce merrifield

high school. He described this period of his life briefly in his autobiography: “The it would take about a decade until proteinogenic amino acids of high quality would become commercially avail- Depression surely affected my life and influenced my future behavior. Stability able. and security became important, but wealth and social position were not. Steady, After a year performing routine tasks as an animal hard work and conservative approaches to problems seem to have been superim- technician, Bruce decided to go to graduate school, so he returned to UCLA in 1944 and began his thesis research posed on whatever genetic traits were present.” under Dunn. At that time, microbiological proce- s a young boy, Bruce enjoyed all sorts of dures were used to assay many commonly occurring things, including playing with his chem- substances. Much of Bruce’s thesis project seems to have istry set and learning photography, devel- been of microbiological ; however, his graduate oping his own film, and making prints. project provided him with unique experiences working AHis favorite subjects in high school were chemistry, with biopolymers and their building blocks. physics, and astronomy, and he had a special inclination More specifically, the work involved substances toward experimental activities. essential for the growth of various Lactobacillus species, 3 He graduated from Montebello High School, in with the aim to apply these bacteria for quantitative East Los Angeles, in 1939. From there, he first went to analysis, especially of amino acids. That was an area Pasadena Junior College for two years and then went to in which his mentor had pioneered, and Bruce, indeed, the University of California, Los Angeles (UCLA), where succeeded in determining the proline content in two he enrolled as a chemistry major. In the university’s new proteins. (It should be pointed out that at this time, chemistry department, he had outstanding teachers. no protein structure had yet been characterized at the Bruce recalled afterward that he especially enjoyed molecular level, but the characterization of insulin courses in qualitative organic analysis and . would be completed soon.) In addition, Bruce analyzed He received his bachelor of science degree in 1943. the occurrence of pyrimidine bases in yeast and sperm His undergraduate biochemistry teacher, Dr. Max DNA. For this work, he received his PhD in 1949. Dunn, was an expert on amino acids and introduced Bruce was offered a fellowship with D.W. Woolley Bruce to the topic. At this time, amino acids were still at the Rockefeller Institute for Medical Research, in New rather exotic compounds. Two of those occurring in York City, shortly before graduation. Woolley was known proteins were not isolated until after World War I, and for his wide interests in biologically active chemical bruce merrifield

compounds such as vitamins, growth factors, antime- the pentapeptide. Using conventional techniques, he was tabolites, and neurotransmitters. able to prepare a sample with the same physical proper- Before moving to New York, Bruce married Eliza- ties and, most important, identical growth-promoting beth “Libby” Furlong, his girlfriend of many years who activity. Other peptides that he subsequently examined, had also been a graduate student at UCLA. including oxytocin/vasopressin intermediates, led him At Rockefeller, Bruce was assigned to a project to the conclusion that those containing either serine or dealing with a new bacterial growth factor called strepo- cysteine were particularly active, whereas the remaining genin that Woolley had discovered and now wanted part could vary considerably, as demonstrated with a few to characterize more fully. The project was to last for additional peptides he made. The threonine analog of several years. Initially, it involved working with a variety the pentapeptide was found to competitively inhibit its of substances, but then it gravitated towards working strepogenin activity. with partial hydrolysates of proteins that had to be puri- y 1950, the methods for peptide synthesis fied and analyzed before they could be assayed. had undergone only moderate changes since For these purposes, unique methods had recently the days of , who pioneered the B 5 been developed by researchers who worked in the same field. Starting from amino acids with complementary building as Woolley’s laboratory at Rockefeller—for protecting groups at their amino and carboxyl functions, example, counter-current distribution by Lyman Craig selective peptide bond formation could be achieved and and amino acid analysis by William Stein and Stanford the product purified, usually by crystallization. This was Moore. Bruce learned and successfully applied their new a slow process with high loss of product, and one that methods to his own project. often failed. To open the way to elongation of the chain, At about this time, Sanger presented the primary selective removal of one protecting group was required in structure of insulin, and from this hormone, well-defined order to allow a second cycle to be initiated in an analo- fragments could be prepared and investigated. In partic- gous way. After a desired number of such cycles, finally all ular, two peptides from its B-chain, a pentapeptide and other protecting groups were removed to give the corre- a heptapeptide that both contain serine, were studied sponding free peptide. because of their high strepogenin activity. Some improvements in this process were intro- To prove that the activity was real and not due to duced during the 1930s, such as the reductive cleavage an artefact or impurity, Bruce embarked on synthesis of of protecting groups by sodium in liquid ammonia. The The early phase of this bruce merrifield is not very well known, and was only briefly described in Bruce’s autobiography. In the versatile benzyloxycarbonyl (Z) group, which allowed Bruce presented his ideas to Woolley, who realized 2001 filmPeptide and and his coworkers to synthesize the their potential and encouraged him to go ahead with the Protein Synthesis: Ori- nonapeptides oxytocin and vasopressin, as well as several project and start developing it. gin and Development, analogs, should also be mentioned. This outstanding The early phase of this is not very well known, and Bruce said he initially work was performed at about the time when Bruce was only briefly described in Bruce’s autobiography. In believed that it would entered the field of peptide synthesis, and it took place the 2001 filmPeptide and Protein Synthesis: Origin and Devel- take three months to across the street from his laboratory. opment, Bruce said he initially believed that it would take develop the proof of After having practiced peptide synthesis for a few three months to develop the proof of concept for the concept for the new years on his own within the strepogenin project, Bruce new procedure, but instead it took three years. A large procedure, but instead began thinking of an alternative, more broadly appli- number of problems of both chemical and practical it took three years. cable strategy that could also be applied to large targets. nature had to be solved, and a multitude of experimental In May 1959, he wrote down a few pages in his labora- parameters needed to be optimized. tory notebook, from which the initial lines below are As a supporting medium for chemical synthesis, quoted: Bruce originally tried cellulose powder. His idea was to 7 A New Approach to the Continuous, Stepwise Synthesis of esterify the first amino acid to the cellulose OH-groups Peptides. and simultaneously exclude the amino acid from under- going further reaction via its carboxyl function, but his There is a need for a rapid, quantitative, automatic method for attempts in that direction failed. Polyvinyl alcohol and the synthesis of long chain peptides. A possible approach may polyacrylamide were also attempted before polystyrene be the use of chromatographic columns where the peptide is came into focus and was more closely investigated. attached to the polymeric packing and added to by an activated Polystyrene functionalized with sulfonic acid amino acid, followed by removal of the protecting group and groups had been applied by others for ion exchange, but with repetition of the process until the desired peptide is built Bruce could not modify such polymers in order to be up. Finally the peptide must be removed from the supporting useful in this context. However, unfunctionalized polysty- medium. rene, crosslinked with different amounts of divinylben- zene, was also available and had promising mechanical properties. This option was especially appealing as, with only a low degree of crosslinks, the polymer swelled bruce merrifield

greatly in organic solvents. It could also easily be deriva- y the middle of 1961, most of the required tized to provide a handle to attach the first amino acid exploratory work involving the choice and to the polymer as a kind of benzyl ester. Esters were and B separate optimal handling of the solid support, still are reliable protecting groups for carboxyl groups, so temporary protection of the amino group, and the this new approach marked an important step forward in peptide coupling reagent, had been made and efforts the synthesis of proteins. could be directed towards an efficient integration of the Before 1960, the choice of amino-protecting gained experiences to make peptides. Resin esterified, as groups and coupling agents for peptide synthesis was indicated previously, was loaded into a specially designed not very large. Among the few protecting groups avail- reaction vessel equipped with a sintered glass filter and able, Z was not quite optimal because it requires rather a stopcock to allow washing and filtering by suction. strong acidic conditions to be cleaved and a benzyl ester All subsequent synthetic steps involving the resin were is not completely stable under such conditions, resulting performed continuously in this vessel, thereby elimi- in some loss of the amino acid on the polystyrene. nating manipulative losses of material. However, after trying a more acid-labile alternative, The resulting cycle included acidic deprotection 9 Bruce decided to avoid new complications and go ahead of the temporary amino-protecting group of the amino with the original choice. Thus, the first Z-protected acid esterified to the resin; washing; deprotonation to amino acid was esterified to derivatized polystyrene, liberate a free amino function; washing; addition of the whereupon Z was cleaved off with acid and the proton- next protected amino acid and DCCI for coupling to ated amino function neutralized with tertiary amine to the just-liberated reactive site; and finally, after DCCI be ready for elongation with the next Z-amino acid. coupling, another washing, with all reagents in excess to To form peptide bonds, Bruce originally had guarantee the various steps proceeded essentially quanti- active esters in mind, but even when used in consider- tatively. Then, new cycles involving the same steps could able excess, they were not reactive enough to couple to be carried out successively until the chain of the desired all free amino groups. Fortunately, another suitable and peptide target was completed. powerful agent, dicyclohexylcarbodiimide (DCCI), had For the first publication describing his new proce- then become available, allowing the chain-elongation dure, Bruce chose to make a tetrapeptide. This required cycle to dipeptide to be completed. a few additional innovations, such as stabilization of the ester bond to the resin, repetition of the coupling step, Bruce’s publica- bruce merrifield tion of this process established solid phase peptide synthesis (SPPS) as a promising and blocking of unreacted amino functions by acetyla- peptide chain to be increased to two residues per day. alternative to the older tion. The completed peptide was cleaved from the resin Starting from resin loaded with the COOH-terminal methods of peptide by saponification. Then, the crude product was carefully amino acid, formation of eight peptide bonds, cleavage synthesis. Although the purified and simultaneously characterized with respect of the product from the support and purification to underlying principles to the presence of all theoretically possible peptidic side biologically fully active bradykinin could then be accom- were questioned by a products, later called deletion peptides. This last step plished in eight working days. few members of the involved using ion-exchange chromatography, where- The rate could be increased to about six residues peptide community, the upon the desired peptide was obtained, pure by several per day using an automatic synthesizer, which was paper attracted enthu- criteria and identical with authentic reference material. constructed in close cooperation with John Stewart, also siastic attention among Bruce’s publication of this process established solid a coworker of Woolley. Subsequently, Stewart made worldwide phase peptide synthesis (SPPS) as a promising alternative a very large number of bradykinin analogs and other and was soon followed to the older methods of peptide synthesis. Although the peptides by automated synthesis. by even more spec- underlying principles were questioned by a few members In addition to the reaction vessel, the automatic tacular work. of the peptide community, the paper attracted enthusi- synthesizer prototype was composed of components to 11 astic attention among chemists worldwide and was soon store, select, and transfer the required reagents, and a followed by even more spectacular work. A very large stepping drum programmer controlled all of the steps. number of the more than 300 papers that Bruce eventu- Although this instrument was described in detail, within ally published dealt with developments and applications a few years, less robust commercial models appeared on of SPPS, of which only a few can be commented on the market. Some of them, unfortunately, discredited here. SPPS severely. Shortly after, the tetrapeptide paper was followed Until around 1965, SPPS was a one-man show, by publications dealing with the synthesis of the nine- but within the next couple of years, Bruce was joined residue peptide bradykinin. This work featured applica- in his efforts by both guest investigators and students, tion of the newly developed t-butoxycarbonyl (Boc), allowing work to expand into the angiotensin, insulin, instead of Z, and incorporation of side-chain-protected oxytocin, and peptide antibiotic fields. With the arrival three-functional amino acids, as well as a generally in 1967 of Bruce’s first postdoctoral fellow, Bernd Gutte optimized technique, allowing the elongation rate of the from Germany, the ambition was raised to the synthesis bruce merrifield

of a protein with enzymatic activity, ribonuclease A, Bruce, on the same page of the report, summa- composed of no less than 124 amino acids. rized the work as follows: “During the past year this The completion of this landmark project required laboratory has continued to concentrate on the solid an incredible 369 chemical reactions and 11,931 steps phase methods for the chemical synthesis of peptides. by the automatic synthesizer, without isolation of any Our most exciting advance was the synthesis of ribo- intermediates. After rigorous purification, it provided a nuclease.” product with, substrate specificity, analytical and kinetic Parallel with more work being done with SPPS, profiles in very close agreement with those of the natural it also became evident that there were side-reactions protein, as judged from amino acid analysis, enzyme and other imperfections that occasionally gave rise to digestion, antibody neutralization, peptide mapping of experimental difficulties, and that, therefore, further a tryptic digest, paper electrophoresis, gel filtration, and improvements in methodology were required. Begin- ion-exchange chromatography. Furthermore, it exhibited ning around 1970, Bruce systematically reinvestigated a specific activity of 78 percent towards yeast RNA. essentially every component and step in his procedure This work furnished experimental support for with the aim to improve it, from the ester link to the 13 Anfinsen’s assertion that the primary structure of a resin, to the relative stabilities of temporary amino- and protein determines its tertiary structure, and it drew side-chain-protecting groups, the coupling efficiency and attention to SPPS from a wider audience than before, its measuring and monitoring, and the final cleavage of including the world of protein chemists and enzymolo- the peptide chain from the support with hydrofluoric gists. Thus, Moore and Stein wrote in the Rockefeller acid (HF). Quantitative measurements and criteria were University Annual Report for 1968–1969, page 18: required and often became routine. These efforts further “Enzyme chemistry in 1969 took a major step forward consolidated the procedure. with the synthesis of bovine pancreatic ribonuclease. The loss of peptide from the original resin, which The chemical structure of the protein had been eluci- occurred also on cleavage of Boc with acid, totalled 83 dated in the preceding decade by and percent, corresponding to an average of 1.4 percent William Stein and their associates on the fifth floor of per step in the ribonuclease synthesis. To avoid this, the Flexner Hall; the chemical synthesis of the enzyme by Pam-resin was introduced. In addition to exhibiting Bruce Merrifield and Bernd Gutte on the fourth floor of about 100 times greater acid stability, it was found to Flexner Hall brought this subject full circle.” bruce merrifield

completely eliminate another problem encountered useful in most instances in this context, cases of severe earlier: the undesirable formation of a peptide family damage by this reagent gradually accumulated. As a with permanently blocked amino functions, which was consequence, an alternative milder SN2 HF procedure generally very difficult to remove from the desired target was developed in the Merrifield laboratory that could be in the final purification step. applied separately or as a first step in combination with Various handles on the polymer, allowing cleavage the earlier method. under very mild acidic conditions or at alternative sites In possession of the improved SPPS methodology, after the peptide chain had been completed, were also Bruce then returned to synthesis of biologically active introduced and increased the scope of SPPS. After puri- peptides on an extended scale. With different aims in fication, large segments prepared on resins such as those mind, a very large number of peptides, such as glucagon, mentioned can be used to make provision for protein gastrin, cecropin, melittin and melittin hybrids, thymo- synthesis by chemical ligation. sins, and immunoglobulin fragments (most of them Further attention was given to the properties of composed of less than 50 amino acids) were successfully the protecting groups used. This was a field that, in the synthesized. 15 meantime, had grown immensely. The selectivity of Particular attention was paid to the antibacterial benzyl-based protection of side-chain OH-, COOH-, cecropin and to the 29-residue hormone glucagon that, and NH2 groups to acid was improved, resulting in together with insulin, is responsible for the regulation reduced amounts of subsequent premature cleavage and of the glucose level in blood. After a reliable synthesis subsequent side-reactions at these sites. for glucagon had been worked out, within a project In model studies, the efficiency of various coupling spanning more than ten years, a very large number conditions was investigated and, using mass spectros- of glucagon analogs were systematically made. In this copy, it could be shown that the amount of deletion and way, a detailed knowledge of the influence of several insertion peptides could be reduced to a remarkable individual amino acids on the hormonal activity was 0.03 to 0.04 percent under optimal conditions with the determined. This work also lead to increasingly powerful Pam-resin, corresponding to a stepwise yield of more glucagon antagonists that were the prerequisite to further than 99.9 percent. For the final cleavage of the product investigations of pharmacological and biological nature, from the resin, liquid HF had been introduced early beautifully exemplifying the power of SPPS in structure- and applied in the synthesis of ribonuclease. Although activity studies of peptide hormones. bruce merrifield

n 1982, Bruce and his coworkers were the first to synthesize an antibacterial cecropin peptide, I which then opened the field to systematic char- acterization of this class of compounds. Cecropin A is a 37-residue peptide of insect origin that belongs to a group of closely related substances involved in the immunological response to bacterial infections. After the structures of the major cecropins had been confirmed by synthesis, a few analogs were prepared and examined for antibacterial activity against various pathogenic bacteria. As an additional feature, the all-D-cecropin A was synthesized and found to generate not only the same electrical conductivity as its enantiomer in lipid bilayers, but also equal activity against representative bacteria. Nobel laureates in chemistry posed for this group photo after the 1991 Nobel Lectures. From left 17 Cecropins become highly helical in nonpolar media. are: , , Yuan T. Lee, , , These and analogous results with enantiomers of two Bruce Merrifield, Richard R. Ernst, Herbert C. Brown, Dudley R. Herschbach. At far right is phys- additional antibiotics allowed Bruce to conclude that ics laureate Hans G. Dehmelt, who was absent when the physics photo was taken. these peptides exert their action by formation of ion- Photo by Felix Aeberli, Swiss Illustrated, courtesy AIP Emilio Segre Visual Archives, channels in cell membranes without interaction with Physics Today Collection chiral species like enzymes or receptors. in the pharmaceutical industry. Early on, solid-phase Today, the strongly increasing interest in all sorts methods impacted nucleic acid synthesis and, more of biologically active peptides requires efficient methods recently, organic chemistry in general (SPOC). Last, but for their synthesis and further studies. Since about 1975, not least, it should be pointed out that SPPS allows an work concerned with peptide synthesis has increasingly individual scientist to conveniently prepare peptides that been based on application of SPPS, especially work on a formerly required large teams of highly trained peptide moderate scale aimed at total synthesis, structure-activity chemists working over long periods of time. studies, and the provision of material for biological inves- Bruce received many awards and honors, tigations, but also work on a considerably larger scale including the Lasker Basic Medical Research Award, the bruce merrifield

Gairdner International Award, the American Chemical to, a role model of a scientist, and a key player in the Society Award for Creative Work in Synthetic Organic golden age of peptide chemistry. Chemistry, the American Peptide Society Alan E. Pierce Award, and the European Peptide Society Josef Author’s note: I was an early guest investigator with Bruce Rudinger Memorial Lecture Award, and he was elected Merrifield and stayed in close contact with him thereafter. I as a member of the National Academy of Sciences in thank David Eaker, a former colleague; Alexander R. Mitchell, 1972. He was awarded the in a former post doc; and Svetlana Mojsov, a former graduate 1984 “for his development of methodology for chemical student of Bruce Merrifield, for generous information and synthesis on a solid matrix.” Besides these accomplish- comments. My final and rather special thanks to Mrs. Elizabeth ments, he received fifteen honorary doctoral degrees. Merrifield for the privilege to author this memoir. Bruce was survived by his wife Elizabeth and their six children—Nancy, Jim, Betsy, Cathy, Laurie, and Sally—as well as sixteen grandchildren. For more details about Bruce’s personal and 19 scientific life and his interactions with members of his research group at the and colleagues around the world, the reader of this memoir is referred to his autobiography, Life During a Golden Age of Peptide Chemistry (American Chemical Society, 1993), and the special issue in 2008 of the journal Peptide Science (volume 90, pages 153-477), which was dedicated to his memory and contains his complete bibliography. Bruce Merrifield is remembered as an unassuming, friendly, and stoic man, a generous and inspiring mentor and colleague, and an optimistic and painstaking scien- tist who had great intuition. He was a person to look up bruce merrifield

SELECTED BIBLIOGRAPHY SELECTED BIBLIOGRAPHY

1949 1966 With M. S. Dunn and L. E. McClure. The determination of proline in protein With A. Marglin. The synthesis of bovine insulin by the solid phase method. hydrolysates with Lactobacillus brevis. J. Biol. Chem. 179:11–18. J. Am. Chem. Soc. 88:5051–5052. With J. M. Stewart and N. Jernberg. Instrument for automated synthesis of 1950 peptides. Anal. Chem. 38:1905–1914. With M. S. Dunn. The microbiological determination of pyrimidines with Lactobacilli. J. Biol. Chem. 186:331–341. 1968 With H. Takashima and V. du Vigneaud. The synthesis of deamino-oxytocin 1952 by the solid phase method. J. Am. Chem. Soc. 90:1323–1325. With D. W. Woolley. The structure and microbiological activity of some dinucle- otides isolated from yeast ribonucleic acid. J. Biol. Chem. 197:521–537. 1969 With B. Gutte. The total synthesis of an enzyme with ribonuclease A activity. 1956 J. Am. Chem. Soc. 91:501–502. With D. W. Woolley. The synthesis of L-seryl-L-histidyl-L-leucyl-L-valyl-L- With B. F. Gisin and D. C. Tosteson. Solid-phase synthesis of the cyclododec- glutamic acid, a peptide with strepogenin activity. J. Am. Chem. Soc. adepsipeptide valinomycin. J. Am. Chem. Soc. 91:2691–2695. 78:4646–4649. 1971 1962 With B. Gutte. The synthesis of ribonuclease A. J. Biol. Chem. 246:1922–1941. 21 Peptide synthesis on a solid polymer. Fed. Proc. 21:412. 1973 1963 With B. W. Erickson. Acid stability of several benzylic protecting groups used Solid phase peptide synthesis. I. The synthesis of a tetrapeptide. J. Am. Chem. Soc. in solid-phase peptide synthesis. Rearrangement of O-benzyltyrosine to 85:2149–2154. 3-benzyltyrosine. J. Am. Chem. Soc. 95:3750–3756.

1964 1974 Solid-phase peptide synthesis. III. An improved synthesis of bradykinin. Biochem- With A. R. Mitchell and J. E. Clarke. Detection and prevention of urethane istry 3:1385–1390. acylation during solid-phase peptide synthesis by anhydride methods. J. Org. Chem. 39:660–668. 1965 Automated synthesis of peptides. Science 150:178–185. 1975 With G. R. Marshall. The synthesis of angiotensins by the solid-phase method. With R. S. Feinberg. Modification of peptides containing glutamic acid by Biochemistry 4:2394–2401. hydrogen fluoride-anisole mixtures.γ –Acylation of anisole or glutamyl nitrogen. J. Am. Chem. Soc. 97:3485–3496. bruce merrifield

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1976 1982 With A. R. Mitchell, B. W. Erickson, M. N. Ryabtsev, and R. S. Hodges. tert- With L. D. Vizioli and H. G. Boman. Synthesis of the antibacterial peptide Butoxycarbonylaminoacyl-4-(oxymethyl)phenylacetamidomethyl-resin, a cecropin A(1-33). Biochemistry 21:5020–5031. more acid-resistant support for solid-phase peptide synthesis. J. Am. Chem. With R. D. DiMarchi, J. P. Tam, and S. B. H. Kent. Weak acid-catalyzed pyrrol- Soc. 98:7357–7362. idone carboxylic acid formation from glutamine during solid-phase peptide synthesis. Minimization by rapid coupling. Int. J. Peptide Protein Res. 19:88–93. 1977 With G. Barany. A new amino protecting group removable by reduction. Chem- 1983

istry of the dithiasuccinoyl (Dts) function. J. Am. Chem. Soc. 99:7363–7365. With J. P. Tam and W. F. Heath. SN2 deprotection of synthetic peptides with a low concentration of HF in dimethyl sulphide: evidence and application in 1978 peptide synthesis. J. Am. Chem. Soc. 105:6442–6455. With A. R. Mitchell, S. B. H. Kent, and M. Engelhard. A new synthetic route to tert-butyloxycarbonylaminoacyl-4-(oxymethyl)phenylacetamidomethyl- 1984 resin, an improved support for solid-phase peptide synthesis. J. Org. Chem. With V.K. Sarin, S. B. H. Kent and A. R. Mitchell. A general approach to the 43:2845–2852. quantitation of synthetic efficiency in solid-phase peptide synthesis as a function of chain length. J. Am. Chem. Soc. 106:7845–7850. 1979 1986 With S. B. H. Kent, A. R. Mitchell, and M. Engelhard. Mechanisms and 23 prevention of trifluoroacetylation in solid-phase peptide synthesis. Solid phase synthesis. Science 232:341–347. Proc. Natl. Acad. Sci. U.S.A. 76:2180–2184. With J.P. Tam and W. F. Heath. Mechanisms for the removal of benzyl protecting groups in synthetic peptides by trifluoromethanesulfonic acid- 1980 trifluoroacetic acid-dimethyl sulphide. J. Am. Chem. Soc. 108:5242–5251. With V. K. Sarin and S. B. H. Kent. Properties of swollen polymer networks. With W. F. Heath. A synthetic approach to structure-function relationships in Solvation and swelling of peptide-containing resins in solid-phase peptide the murine epidermal growth factor molecule. Proc. Natl. Acad. Sci. U.S.A. synthesis. J. Am. Chem. Soc. 102:5463–5470. 83:6367–6371.

1981 1987 With S. Mojsov. Solid phase synthesis of crystalline glucagon. Biochemistry With C. G. Unson, D. Andreu, and E. M. Gurzenda. Synthetic peptide antago- 20:2950–2956. nists of glucagon. Proc. Natl. Acad. Sci. U.S.A. 84:4083–4087. With V. K. Sarin, S. B. H. Kent, and J. P. Tam. Quantitative monitoring of solid- phase peptide synthesis by the ninhydrin reaction. Anal. Biochem. 117:147–157. SELECTED BIBLIOGRAPHY

1988 With E. Kaiser, Sr., J. P. Tam and T.M. Kubiak. Chlorotrimethylsilane-phenol as a mild deprotection reagent for the tert-butyl based protecting groups in peptide synthesis. Tetrahedron Lett. 29:303–306. With J. Singer and B. T. Chait. Mass spectrometric evaluation of synthetic peptides for deletions and insertions. Anal. Biochem. 174:399–414.

1990 With D. Wade, A. Boman, B. Wåhlin, C. M. Drain, D. Andreu, and H. G. Boman. All D-amino acid-containing channel-forming antibiotic peptides. Proc. Natl. Acad. Sci. U.S.A. 87:4761–4765.

1993 Life during a golden age of peptide chemistry. The concept and development of solid-phase peptide synthesis. Washington, DC: American Chemical Society.

1995 With E. L. Merrifield, S. A. Mitchell, J. Ubach, H. G. Boman, and D. Andreu. D-enantiomers of 15-residue cecropin A-melittin hybrids. Int. J. Peptide Protein Res. 46:214–220.

1996 With P. Juvvadi and S. Vunnam. Synthetic melittin, its enantio, retro, and retroe- nantio isomers, and selected chimeric analogs: their antibacterial, hemolytic, and lipid bilayer action. J. Am. Chem. Soc. 118:8989–8997.

1999 With F. M. Marassi, S. J. Opella, and P. Juvvadi. Orientation of cecropin A helices in phospholipid bilayers determined by solid-state NMR spectros- Published since 1877, Biographical Memoirs are brief biographies of deceased copy. Biophys. J. 77:3152–3155. National Academy of Sciences members, written by those who knew them 2003 or their work. These biographies provide personal and scholarly views of History of protein synthesis. In Houben-Weyl, Synthesis of Peptides and Peptidomi- America’s most distinguished researchers and a biographical history of U.S. metics, M. Goodman, A. Felix, L. Moroder, and C. Toniolo (eds.) E22b:3-41. science. Biographical Memoirs are freely available online at www.nasonline.org/ Stuttgart: Thieme Publishers. memoirs. 26