Authors: Lila M. Gierasch | Fernando Alberício | Claudio Toniolo
Source: Peptide Science (article, 2020-07-01)
Field: Chemistry — Physical and Theoretical Chemistry

Dedicated to a peptide chemist of few words and immense impact: Louis A. Carpino This issue of Peptide Science is a testimonial to brilliance packaged in a nontraditional way. We are honoring the contributions of Louis A. Carpino (“Lou” to many of us or “Louie” to a few), who passed away in January 2019 at the age of 91. Lou Carpino's contributions to peptide chemistry specifically, and organic chemistry more broadly, over the course of his career are truly stunning as summarized in the articles in this collection. Yet he pursued his work quietly and on his own terms, eschewing the usual accolades and attention that would come to one whose work is so influential. The compilation of papers in this issue constitutes a testament to the impact of Lou Carpino's research. However, the articles are only representative as there is little in the peptide chemistry field that has not been enabled and touched by the genius and chemical virtuosity of Lou Carpino. In addition to the research articles collected here, we have also gathered brief “In Memoriam” pieces from a number of Lou's colleagues. These pieces paint a fuller picture of Lou Carpino the mentor, Lou Carpino the teacher, and—to a small extent—Lou Carpino the father. A master of compartmentalization, Lou Carpino was for each of us only one thread of a complex fabric. We hope that this issue will give a better picture of the full fabric of Lou Carpino and, in so doing, reflect the masterful chemist and multifaceted human being he was. The field of peptide chemistry is richer for his stellar career-long contributions and poorer for the passing of this gentle and passionate man. Lou Carpino was born in 1927 in Des Moines, IA (U.S.A.). He took his academic position at the University of Massachusetts at Amherst in 1954 after completing his doctoral work at the University of Illinois with Harold R. Snyder and a short postdoctoral stint at the University of South Carolina with DeLos F. Detar. He published his first independent paper based on work at UMass Amherst in 1956, and many other single-author papers followed shortly. These were harbingers of the abundant and significant publications emanating from Lou Carpino's lab. Lou Carpino remained at UMass Amherst for his entire 65-year career, active until shortly before his death. We begin this issue with editorial pieces reflecting several aspects of Lou Carpino's career. First, we reprint an obituary written by Lou Carpino's longtime collaborator, Michael Beyermann. Lou Carpino worked for many years in a three-way collaboration with Michael Beyermann and Michael Bienert, and was also joined in this collaboration by Holger Wenschuh, then a graduate student. We include touching comments about the scientific partnership from Holger Wenschuh. Following these comments is a short piece by one of us (Fernando Albericio) putting Lou Carpino's contributions to peptide chemistry in perspective. Additional pieces from Ayman El-Faham, Khaled Nasr and Salvatore “Sal” Triolo, all of whom were trainees with Lou Carpino, and Peter Lillya, a longtime faculty colleague of Lou Carpino's at UMass, paint a picture of Carpino the teacher and mentor. Another of the co-editors (Lila Gierasch) offers additional thoughts on Lou Carpino as a colleague and friend, including brief comments about his family. A detailed commentary from the third co-editor (Claudio Toniolo) emphasizing Lou's scientifically very productive 6-month stay at his Institute in Padua in the late 1960s can be found in the Introduction of a paper in this special issue (C. Toniolo, A. Moretto, F. Formaggio, Q. B. Broxterman and M. Crisma, Pept. Sci. 2020, 112, e24153). Following the introductory material to this issue are several articles contributed by members of the peptide chemistry community. These illustrate how greatly this community values the work of Lou Carpino and how peptide chemistry is enabled by the landmark achievements Carpino made to protecting groups and coupling reagents. Please read the articles and realize as you do how much of the work in the peptide field, currently and over the past 50 years, could not have happened without the contributions of Lou Carpino. We hope that this issue of Peptide Science stands as a testament to a spectacular career and that, more importantly perhaps, it reveals and shines light on a scientist who chose to stay away from the limelight while he passionately pursued his research. We also hope that you will get to know Lou Carpino better than you may have had the chance to while he was alive, sadly. Lila M. Gierasch1 Fernando Albericio2,3 Claudio Toniolo4 1Departments of Biochemistry & Molecular Biology and Chemistry, University of Massachusetts, Amherst, Amherst, U.S.A. 2School of Chemistry, University of KwaZulu-Natal Durban, South Africa 3Department of Organic Chemistry, University of Barcelona Barcelona, Spain 4Department of Chemistry, University of Padua and Institute of Biomolecular Chemistry, Padua Unit, CNR, Padua, Italy Special Collections & University Archives The University of Massachusetts Amherst A treasure trove of material from Louis A. Carpino's career has been archived in the Special Collections of the University of Massachusetts Amherst library archives. It is available as an invaluable resource chronicling the intellectual path through peptide chemistry followed by this giant of our field. Among the items that have been archived are over 20 notebooks kept by Louis Carpino from the time he started his research. They are written in his own hand and describe his thinking and creative experimental design, which led to the products he has offered the field. We urge anyone interested to avail themselves of this material through a visit to the University campus in Amherst. Please make a virtual visit to the archives in the meantime at: http://scua.library.umass.edu/umarmot/carpino-louis-a/. Louis A. Carpino (1927-2019) Dr Louis A. Carpino (91) passed away at home in Amherst, MA, on Friday, January 4, 2019. He was a Professor Emeritus of Organic Chemistry at the University of Massachusetts Amherst. Dr L. A. Carpino was a pioneer in the development of amino-protecting groups and coupling reagents for use in the synthesis of biologically active materials such as pharmaceuticals, polynucleotides, peptide nucleic acids (PNAs), peptides, and small proteins. His first significant contribution was the development of the tert-butyloxycarbonyl (Boc) group as a protecting group for amino acids.[1] With its much greater acid lability than the benzyloxycarbonyl-(Z) protecting group, the Boc group facilitated the synthesis of peptides. The discovery of the Boc group enabled a new strategy in the synthesis of peptides. The Boc strategy combined the use of a Boc group as a temporary Nα-amino protecting group with the much more acid-stable, permanent side chain benzyl-based protecting groups. Boc amino acids are mostly crystalline, easily prepared using a number of synthetic methods, and stable even at room temperature. In general, activation and coupling of Boc amino acids to the N-terminus of growing peptides is carried out with no side reaction. Because of the rapid removal of Boc groups by TFA/DCM, HCl/dioxane or other acids, Boc amino acids were found to be particularly suited for the development of and use in solid-phase peptide synthesis (SPPS) created by R. B. Merrifield in 1963.[2] Boc protection of amino acids has been highly successful in the syntheses of a limited number of proteins and is compatible with automated machines and for parallel synthesis of a large number of short peptides (on pins or in “tea bags”). Dr Carpino's next major accomplishment addressed a weak point in the use of Boc-protecting groups, especially in the presence of other acid labile-protecting groups. While the temporary Boc group can be removed under much milder conditions than the permanent groups, the latter groups are not completely stable following repeated Boc removals. Thus, partial side-chain deprotection during the synthesis of long peptides in SPPS may yield undesired side products and complex crude products. Moreover, the final deprotection of side chain functionalities, as well as cleavage of peptide from resin in the Boc strategy, necessitates the use of harsh conditions, such as liquid hydrogen fluoride, which may lead to modification/degradation of the product by side reactions, including aspartimide formation, N/O-shift, etc. Dr Carpino developed a completely orthogonal-protecting system using the 9-fluorenylmethoxycarbonyl (Fmoc) group.[3] The Fmoc-protecting group is removed rapidly and efficiently by proton abstraction via secondary amines, conditions that do not affect acid-labile tert-butyl-type protecting groups placed on the side chains. Moreover, the Fmoc group is completely stable under the conditions used for removal of the resin linkers. Fmoc amino acids are generally easy to prepare in crystalline form and stable during storage. Fmoc amino acids can be highly activated and efficiently coupled without side reactions, and their high ultraviolet (UV) absorption makes it possible to monitor their incorporation during the assembly of the peptide. By using the secondary amines such as piperidine to remove the Fmoc group, the dibenzofulvene formed is trapped on the spot, minimizing possible side reactions. The Fmoc-based SPPS is currently the method of choice for the chemical synthesis of peptides. Dr Carpino developed several other protecting groups that have also found widespread application. The benzothiophenesulfone-2-methyloxycarbonyl (Bsmoc)-protecting group is an example of a large new class of protecting groups that can be removed under the very mild conditions of Michael addition.[4] The 2,2,4,6,7-pentamethyldihydrobenzofuran-5-sulfonyl group (Pbf) was developed for side chain protection of arginine and is easily removed by trifluoroacetic acid (TFA). Pbf can be used efficiently in the synthesis of arginine-containing peptides.[5] The icyclopropylmethyl (Dcpm) group is used in polypeptide syntheses when it is necessary to minimize peptide insolubility and aggregation.[6] In the case of threonine/serine-containing peptides, an alternative route to avoid aggregation problems involves the synthesis of the corresponding O-acyl-peptide isomers and subsequent rearrangement back to regular peptides after synthesis and purification.[7] In addition to his seminal contributions around amino-protecting groups, Dr Carpino developed several new types of coupling reagents based on acid halides and 1-hydroxy-7-azabenzotriazole that are far more efficient than classical reagents because of their speed, generality, and control of epimerization. Fmoc-amino acid fluorides have facilitated efficient SPPS of peptaibols, such as alamethicin, the synthesis of which was previously reported via a laborious solution-phase route.[8] The more reactive Fmoc-amino acid chlorides, although being less stable than the corresponding fluorides, enable the coupling of extremely hindered systems, such as N-methyl-amino-iso-butyric acid to amino-iso-butyric acid.[9] Finally, uronium/guanidinium compounds of 1-hydroxy-7-azabenzotriazole (HOAT) are very efficient activation reagents,[10] although their recent use, especially in large-scale syntheses, has been limited for safety reasons. Dr Carpino's contributions over a long and distinguished career have enabled synthetic peptide chemistry as it is widely practiced today. Dr Carpino never stopped thinking about the design of better coupling reagents. In fact, he was working on a new coupling reagent to address some of the safety issues with HOAT over the past year when he passed away. Dr Carpino was born in Des Moines, Iowa, on December 13, 1927. He was a loving husband, father, and grandfather and is survived by his wife, Barbara, and six children. I got to know him as a great teacher, creative director who shared his ideas, attentive listener who showed respect for anyone, a quiet person who shunned the great arena but fostered intense scientific collaboration over many years. He touched the lives of many people—he will be missed. Michael Beyermann Longtime collaborator of Louis Carpino Markkleeberg, Germany [1] L.A. Carpino, J. Am. Chem. Soc. 1957, 79, 4427. [2] R. B. Merrifield, J. Am. Chem. Soc. 1963, 85, 2149. [3] L. A. Carpino, G.Y. Han, J. Am. Chem. Soc. 1970, 92, 5748. [4] L. A. Carpino, M. Ismail, G. A. Truran, E. M. E. Mansour, S. Iguchi, D. Ionescu, A. El-Faham, C. Riemer, R. Warrass, J. Org. Chem. 1999, 64, 4324. [5] L. A. Carpino, H. Shroff, S. A. Triolo, E. M. E. Mansour, Tetrahedron Lett. 1993, 34, 7829. [6] L. A. Carpino, K. Nasr, A. A. Abdel-Maksoud, A. El-Faham, D. Ionescu, P. Henklein, H. Wenschuh, M. Beyermann, E. Krause, M. Bienert, Org. Lett. 2009, 11, 3718. [7] L. A. Carpino, Krause E, C. D. Sferdean, M. Schümann, H. Fabian, M. Bienert, M. Beyermann, Tetrahedron Lett. 2004, 45, 7519. [8] L. A. Carpino, M. Beyermann, H. Wenschuh, M. Bienert, Acc. Chem. Res. 1996, 29, 68. [9] L. A. Carpino, D. Ionescu, A. El-Faham, P. Henklein, H. Wenschuh, M. Bienert, M. Beyermann, Tetrahedron Lett. 1998, 39, 241. [10] L. A. Carpino, H. Imazumi, A. El-Faham, F. J. Ferrer, C. Zhang, Y. Lee, B. M. Foxman, P. Henklein, C. Hanay, C. Mügge, H. Wenschuh, J. Klose, M. Beyermann, M. Bienert, Angew. Chem. Int. Ed. 2002, 41, 441. Louis A. Carpino (1927-2019) Michael Beyermann J. Pept. Sci. 2019, 25, e3168; John Wiley and Sons, Ltd © 2019 European Peptide Society and John Wiley & Sons, Ltd. Reprinted with permission of Wiley Publishing, Ltd. In 1990, when I started my Ph.D. work at the East-German Institute for Molecular Pharmacology in Berlin, only a few months after the Berlin Wall collapsed, Louis Carpino had an ongoing collaboration with our research team. This was unusual because the beginning of that fruitful relationship dated back to the time when the institute still worked under the strict East German regulations. When Louis and his lovely wife Barbara visited Berlin and the institute early in 1991, he asked whether I could imagine coming to the United States to do part of my thesis work in his laboratories at the University of Massachusetts at Amherst. As a young East German scientist I was of course full of excitement and applied for a grant at the German Academic Exchange Service. Almost nine months later, on January 6, 1992, Louis and Barbara picked me up at Bradley Airport in Hartford, Connecticut, and gave me a ride to Amherst. The following seven months under Louis' mentorship became some of the most influential of my entire academic and business career. Louis A. Carpino's research team in 1992 [Reprinted with permission of the University of Massachusetts Chemistry Department.] We started a variety of projects exploiting the full impact of protected amino acid fluorides for solid phase peptide synthesis, their suitability for incorporating extremely sterically hindered amino acids into peptides, for the assembly of biologically active peptaibols, and more. During that time I witnessed Louis' outstanding ability to manage a multinational scientific team with members from eight countries and varying cultural backgrounds. He was not only always open to new ideas and visions but he actively supported all of us by sharing his profound knowledge of peptide chemistry, including how to structure scientific work in a systematic and goal-oriented way. His capacity to listen with respect for everyone and to motivate every single team member, especially after setbacks, was inspiring and gave me guidance throughout my business career. My first visit to Louis“lab was not the last. Only one year later a grant from Perseptive Biosystems supported by Fernando Albericio brought me back to Louis' lab for another couple of months. This was like coming home. During my academic career—post-docing at the Max Planck Institutes in Germany; joining JERINI AG, a peptide drug company; and finally establishing JPT Peptide Technologies in Berlin—we continued to have an active scientific and personal exchange. Louis and Barabara's annual visits to Berlin were always among the highlights of the year and felt like family reunions. Over the years we were able to co-publish more than 30 peer-reviewed papers, short communications and book chapters. Louis' passing in 2019 was an incredible loss for the peptide science community and me. His countless contributions to modern peptide science laid the foundation for many of the most recent breakthrough applications using thousands of peptides in proteomics and immunotherapy. Every single day we continue to apply and improve the knowledge Louis Carpino created and shared over the decades for the sake of developing advanced therapeutic options for meeting medical needs. Holger Wenschuh Collaborator and visiting scientist with Louis Carpino Managing Director, JPT Peptide Technologies GmbH, 12489 Berlin, Germany My colleague Louis Carpino I met Prof. Carpino when I was appointed Director of Peptide Research at Millipore-Waters near Boston, U.S.A., in 1991. My first relation with him in this position was rather bizarre for two scientists from academia (I was on leave from the University of Barcelona, Spain, where I held a position as Associate Professor, and where I returned in 1994 to continue my academic career). In this regard, we were brought together to negotiate the commercialization rights of the new coupling reagents based on the 7-aza-1-hydroxybenzotriazole (HOAt), the the and the that we had developed the two In Prof. Carpino was interested in Millipore-Waters the commercialization of the protecting to was to Prof. Carpino and the University of Massachusetts were by Research Technologies a based in I first met the Director of on a in MA, and I that he brought me of most of Prof. Carpino's papers to me who this scientist was in the peptide and the of coupling reagents and protecting groups. the of our the director of and that I more about Prof. Carpino and his work than he As you may we an and I hope Prof. Carpino and UMass took away of their with However, there is no that who most from that were the members of the peptide community. to the reagents and such as and and still available to and after intense of several including which later the rights of the with UMass, and finally the family at the acids are no a a more personal I Prof. Carpino a from whom I several In addition to his scientific I had the of meeting his Prof. Ayman and I first started our collaboration under the of Prof. Carpino and then continued it This collaboration has 30 years, and following Prof. Carpino's scientific we developed I also met Dr Fernando who is currently in and with whom I still two of Prof. Carpino's Dr who developed the Pbf group, and Dr Salvatore Triolo, joined my group at Prof. Carpino's were and reflect the of the great Louis Carpino, an peptide chemist can who had a profound impact on the peptide in Chemistry in and R. Merrifield Yet while these scientists made seminal could not be organic and their work not the of peptide In the peptide the two organic were and Louis A. Carpino. The was for the of the more as protecting was that the group the of a organic after of other of the the protecting group would then be the group of Louis Carpino took this one as he that to be a protecting group had to be and that protecting group removal under mild conditions to the of the peptide his career in peptide chemistry, Carpino was or better by under mild conditions, and the of the compounds This the organic of thinking and with that of the who is more interested in the although some have to be As peptide we can our that this was the of Carpino's career because it led to many groups and that have the peptide field in Carpino developed chemistry for the his achievements were rapidly to solid-phase With respect to Peptide two are always and are based on Boc and two protecting groups developed by Carpino. The was published in while the latter about in while Carpino was for protecting groups in mild The Boc group was rapidly by Merrifield for the synthesis of published in his seminal Merrifield used by which to be removed in solid the Fmoc group, in and in the U.S.A. and and in developed a mild SPPS strategy, As in an of this issue of Peptide Carpino's work on protecting groups far Boc and In the field of protecting groups, he much protecting groups. However, his most significant contribution to this field was the development of a new family of protecting groups, which are removed through a Michael addition reaction. The most representative example of this family is to Carpino showed several over but as was reported amino acids have never the The most amino acid side chain to is the of is that the protecting group of choice is 2,2,4,6,7-pentamethyldihydrobenzofuran-5-sulfonyl group developed by is available and is by far the most widely used in of the of the Carpino developed protecting groups based on that for the group and for the side chain of and He the used for the side chain of for with the of and which are the for the of With the the of chain collaboration with the group of his and Beyermann, and in parallel to Professor work in the of and in the peptide chain through the of these amino acids hydrogen the is and the is to the peptide in Carpino was also a in of Peptide and in he developed and solid for the rapid and removal of the Fmoc group in A few years later, he rapid Peptide for the of peptides with up to amino acids based on the use of the for Fmoc The to this strategy is the use of an to remove the from the of dibenzofulvene with the secondary and the As in one of the contributions to this peptide synthesis is as the of protecting groups and coupling reagents. With this in Carpino all peptide In the first part of his scientific career he worked on the development of protecting groups, while in the part he the of coupling the in the of chemistry, we can that Carpino pursued the use of by impact in of and very high he gave a to halides and as a coupling In the case of the which is more compatible with the side chain protecting groups used in the Fmoc he developed the first coupling reagent which in the corresponding acid with even the chemistry, was developed by Carpino as an for the in of the acid using The contribution of Carpino to this field was the development of the chemistry based on 7-aza-1-hydroxybenzotriazole which has been to greatly in solid-phase and chemistry, in of coupling and the corresponding and were These reagents are the most efficient in the solid phase and phase for the of highly hindered These include and amino These reagents have for the in the synthesis of peptides. Carpino showed he had the of presence of the in position of the system had a group on the he was greatly by the of its corresponding the first of this family of compounds was to where the of the was to a In an to the of and Carpino and that reagent was a but were where the the two was through a of the and the He later on to and a This of Louis Carpino's contributions to peptide synthetic chemistry is offered to a genius he was and the of his Fernando Albericio Longtime collaborator of Louis Carpino of Chemistry, University of Durban, South Africa [1] F. E. Chem. 34, [2] C. C. J. M. C. P. G. S. J. Am. Chem. Soc. [3] R. B. Merrifield, J. Am. Chem. Soc. 1963, 85, 2149. [4] L. Chem. [5] L. A. Carpino, Chem. Res. 6, [6] L. A. Carpino, J. Am. Chem. Soc. [7] L. A. Carpino, G. Y. Han, J. Am. Chem. Soc. 1970, 92, 5748. [8] R. B. Merrifield, J. Am. Chem. Soc. [9] J. Int. J. Res. 11, [10] E. H. D. C. J. R. C. B. J. J. Chem. Chem. A. A. El-Faham, F. Pept. Sci. 2020, 112, L. A. Carpino, M. M. Ismail, G. A. Truran, E. M. E. Mansour, S. Iguchi, D. Ionescu, A. El-Faham, C. Riemer, R. Warrass, M. S. J. Am. Chem. Soc. L. A. Carpino, H. Shroff, S. A. Triolo, E. S. M. E. Mansour, H. Wenschuh, F. Tetrahedron Lett. 1993, 34, 7829. L. A. Carpino, H. G. S. E. M. E. Mansour, C. Riemer, R. Warrass, D. G. A. Truran, H. Imazumi, H. Wenschuh, M. Beyermann, M. Bienert, H. Shroff, F. S. A. Triolo, A. Chemist ; Art ; Art history ; Chemistry ; Organic chemistry History and advancements in chemistry ; Various Chemistry Research Topics