Editing Insulin (section)

== History of study ==

=== Discovery ===
In 1869, while studying the structure of the [[pancreas]] under a [[microscope]], [[Paul Langerhans]], a medical student in [[Berlin]], identified some previously unnoticed tissue clumps scattered throughout the bulk of the pancreas.<ref>{{cite journal | vauthors = Sakula A | title = Paul Langerhans (1847-1888): a centenary tribute | journal = Journal of the Royal Society of Medicine | volume = 81 | issue = 7 | pages = 414–5 | date = July 1988 | pmid = 3045317 | pmc = 1291675 | doi = 10.1177/014107688808100718 }}</ref> The function of the "little heaps of cells", later [[eponym|known as]] the ''[[islets of Langerhans]]'', initially remained unknown, but [[Édouard Laguesse]] later suggested they might produce secretions that play a regulatory role in digestion.<ref>{{cite web |url=http://musee.chru-lille.fr/Memoire/Medecins/48029.html |title=Edouard Laguesse (1861–1927) | vauthors = Petit H |language=fr |website=Museum of the Regional Hospital of Lille |access-date=25 July 2018 }}</ref> Paul Langerhans' son, Archibald, also helped to understand this regulatory role.

In 1889, the physician [[Oskar Minkowski]], in collaboration with [[Joseph von Mering]], removed the pancreas from a healthy dog to test its assumed role in digestion. On testing the urine, they found sugar, establishing for the first time a relationship between the pancreas and diabetes. In 1901, another major step was taken by the American physician and scientist [[Eugene Lindsay Opie]], when he isolated the role of the pancreas to the islets of Langerhans: "Diabetes mellitus when the result of a lesion of the pancreas is caused by destruction of the islets of Langerhans and occurs only when these bodies are in part or wholly destroyed".<ref name="Johns Hopkins Hosp. Bull. p.264)">{{cite journal | vauthors = Opie EL | title = Diabetes Mellitus Associated with Hyaline Degeneration of the islands of Langerhans of the Pancreas | journal = Bulletin of the Johns Hopkins Hospital | volume = 12 | issue = 125 | pages = 263–64 | year = 1901 | hdl = 2027/coo.31924069247447 }}</ref><ref name="J. Exp. Med. 5(4)">{{cite journal | vauthors = Opie EL | title = On the Relation of Chronic Interstitial Pancreatitis to the Islands of Langerhans and to Diabetes Mellitus | journal = Journal of Experimental Medicine | volume = 5 | issue = 4 | pages = 397–428 | year = 1901 | doi=10.1084/jem.5.4.397| pmid = 19866952 | pmc = 2118050 }}</ref><ref name="J. Exp. Med. 5(5)">{{cite journal | vauthors = Opie EL | title = The Relation of Diabetes Mellitus to Lesions of the Pancreas. Hyaline Degeneration of the Islands of Langerhans | journal = Journal of Experimental Medicine | volume = 5 | issue = 5 | pages = 527–40 | year = 1901 | doi=10.1084/jem.5.5.527| pmid = 19866956 | pmc = 2118021 }}</ref>

Over the next two decades researchers made several attempts to isolate the islets' secretions. In 1906 [[George Ludwig Zuelzer]] achieved partial success in treating dogs with pancreatic extract, but he was unable to continue his work. Between 1911 and 1912, [[Ernest Lyman Scott|E.L. Scott]] at the [[University of Chicago]] tried aqueous pancreatic extracts and noted "a slight diminution of glycosuria", but was unable to convince his director of his work's value; it was shut down. [[Israel Kleiner (biochemist)|Israel Kleiner]] demonstrated similar effects at [[Rockefeller University]] in 1915, but [[World War I]] interrupted his work and he did not return to it.<ref name="J. Nutrition 92">{{cite journal | author = The American Institute of Nutrition |title=Proceedings of the Thirty-First Annual Meeting of the American Institute of Nutrition | journal = Journal of Nutrition | volume = 92 | issue = 4 | year = 1967 | page = 509 |doi= 10.1093/jn/92.4.507 }}</ref>

In 1916, [[Nicolae Paulescu]] developed an [[aqueous]] [[Pancreas|pancreatic]] extract which, when injected into a [[Diabetes|diabetic]] dog, had a normalizing effect on [[blood sugar]] levels. He had to interrupt his experiments because of [[World War I]], and in 1921 he wrote four papers about his work carried out in [[Bucharest]] and his tests on a diabetic dog. Later that year, he published "Research on the Role of the [[Pancreas]] in Food Assimilation".<ref name="nrjs">{{cite journal | vauthors = Paulesco NC | journal = Archives Internationales de Physiologie |url= https://insulin.library.utoronto.ca/islandora/object/insulin%3AT10137 | title= Recherche sur le rôle du pancréas dans l'assimilation nutritive|volume= 17|pages= 85–109| date=31 August 1921 }}
</ref><ref name="nrps">
{{cite journal | vauthors = Lestradet H | journal = Diabetes & Metabolism | title = Le 75e anniversaire de la découverte de l'insuline | volume = 23 | issue = 1| page = 112 | year = 1997 | url=  http://www.em-consulte.com/en/article/79613 }}
</ref>

The name "insulin" was coined by [[Edward Albert Sharpey-Schafer]] in 1916 for a hypothetical molecule produced by pancreatic islets of Langerhans (Latin ''insula'' for islet or island) that controls glucose metabolism. Unbeknown to Sharpey-Schafer, Jean de Meyer had introduced the very similar word "insuline" in 1909 for the same molecule.<ref>{{Cite journal| vauthors = de Leiva A, Brugués E, de Leiva-Pérez A |date=2011|title=The discovery of insulin: Continued controversies after ninety years|journal=Endocrinología y Nutrición (English Edition)|language=en|volume=58|issue=9|pages=449–456|doi=10.1016/j.endoen.2011.10.001}}</ref><ref>{{cite journal | vauthors = Vecchio I, Tornali C, Bragazzi NL, Martini M | title = The Discovery of Insulin: An Important Milestone in the History of Medicine | journal = Frontiers in Endocrinology | volume = 9 | pages = 613 | date = 23 October 2018 | pmid = 30405529 | pmc = 6205949 | doi = 10.3389/fendo.2018.00613 | doi-access = free }}</ref>

=== Extraction and purification ===
In October 1920, Canadian [[Frederick Banting]] concluded that the digestive secretions that Minkowski had originally studied were breaking down the islet secretion, thereby making it impossible to extract successfully. A surgeon by training, Banting knew that blockages of the pancreatic duct would lead most of the pancreas to atrophy, while leaving the islets of Langerhans intact. He reasoned that a relatively pure extract could be made from the islets once most of the rest of the pancreas was gone. He jotted a note to himself: "Ligate pancreatic ducts of dog. Keep dogs alive till acini degenerate leaving Islets. Try to isolate the internal secretion of these + relieve glycosurea[sic]."<ref>{{cite web|url=https://insulin.library.utoronto.ca/islandora/object/insulin%3AN10002|title=Note dated Oct 31/20 from loose leaf notebook 1920/21| vauthors = Banting FG |date=31 October 1920|website=University of Toronto Libraries}}</ref><ref name="rosenfeld2002">{{cite journal | vauthors = Rosenfeld L | title = Insulin: discovery and controversy | journal = Clinical Chemistry | volume = 48 | issue = 12 | pages = 2270–88 | date = December 2002 | pmid = 12446492 | doi = 10.1093/clinchem/48.12.2270 | doi-access = free }}</ref>
[[File:Charles H. Best and Clark Noble ca. 1920.jpg|thumb|left|[[Charles Best (medical scientist)|Charles Best]] and Clark Noble ca. 1920]]
In the spring of 1921, Banting traveled to [[Toronto]] to explain his idea to [[John Macleod (physiologist)|John Macleod]], Professor of Physiology at the [[University of Toronto]]. Macleod was initially skeptical, since Banting had no background in research and was not familiar with the latest literature, but he agreed to provide lab space for Banting to test out his ideas. Macleod also arranged for two undergraduates to be Banting's lab assistants that summer, but Banting required only one lab assistant. [[Charles Best (medical scientist)|Charles Best]] and Clark Noble flipped a coin; Best won the coin toss and took the first shift. This proved unfortunate for Noble, as Banting kept Best for the entire summer and eventually shared half his Nobel Prize money and credit for the discovery with Best.<ref name="pmid12473641">{{cite journal | vauthors = Wright JR | title = Almost famous: E. Clark Noble, the common thread in the discovery of insulin and vinblastine | journal = CMAJ | volume = 167 | issue = 12 | pages = 1391–96 | date = December 2002 | pmid = 12473641 | pmc = 137361 }}</ref> On 30 July 1921, Banting and Best successfully isolated an extract ("isletin") from the islets of a duct-tied dog and injected it into a diabetic dog, finding that the extract reduced its blood sugar by 40% in 1 hour.<ref name="Krishnamurthy2002">{{cite book | vauthors = Krishnamurthy K | title = Pioneers in scientific discoveries | url = https://books.google.com/books?id=dAXYzzDL_9oC&pg=PA266 | access-date = 26 July 2011 | year = 2002 | publisher = Mittal Publications | isbn = 978-81-7099-844-0 | page=266 }}</ref><ref name="rosenfeld2002"/>

Banting and Best presented their results to Macleod on his return to Toronto in the fall of 1921, but Macleod pointed out flaws with the experimental design, and suggested the experiments be repeated with more dogs and better equipment. He moved Banting and Best into a better laboratory and began paying Banting a salary from his research grants. Several weeks later, the second round of experiments was also a success, and Macleod helped publish their results privately in Toronto that November. Bottlenecked by the time-consuming task of duct-tying dogs and waiting several weeks to extract insulin, Banting hit upon the idea of extracting insulin from the fetal calf pancreas, which had not yet developed digestive glands. By December, they had also succeeded in extracting insulin from the adult cow pancreas. Macleod discontinued all other research in his laboratory to concentrate on the purification of insulin. He invited biochemist [[James Collip]] to help with this task, and the team felt ready for a clinical test within a month.<ref name="rosenfeld2002"/>
[[File:Chart for Elizabeth Hughes (12308739143).jpg|thumb|Chart for Elizabeth Hughes, used to track blood, urine, diet in grams, and dietary prescriptions in grams]]
On 11 January 1922, [[Leonard Thompson (diabetic)|Leonard Thompson]], a 14-year-old diabetic who lay dying at the [[Toronto General Hospital]], was given the first injection of insulin.<ref name="pmid8409364">{{cite journal | vauthors = Bliss M | title = Rewriting medical history: Charles Best and the Banting and Best myth | journal = Journal of the History of Medicine and Allied Sciences | volume = 48 | issue = 3 | pages = 253–74 | date = July 1993 | pmid = 8409364 | doi = 10.1093/jhmas/48.3.253 | url = https://academic.oup.com/jhmas/article-pdf/48/3/253/9838324/253.pdf | doi-access = free }}</ref><ref>{{cite web|url=https://insulin.library.utoronto.ca/islandora/object/insulin%3AC10024|title=Work on diabetes shows progress against disease| work = Toronto Star Weekly|date=14 January 1922| publisher = University of Toronto Libraries}}</ref><ref>{{cite journal | vauthors = Fletcher AA | title = Early clinical experiences with insulin | journal = Canadian Medical Association Journal | volume = 87 | pages = 1052–5 | date = November 1962 | issue = 20 | pmid = 13945508 | pmc = 1849803 | url = https://insulin.library.utoronto.ca/islandora/object/insulin%3AT10053 }}</ref><ref>{{cite web|url=https://insulin.library.utoronto.ca/islandora/object/insulin%3AM10015|title=Patient records for Leonard Thompson| vauthors = Banting FG |date=Dec 1921 – Jan 1922|website=University of Toronto Libraries}}</ref> However, the extract was so impure that Thompson had a severe [[anaphylaxis|allergic reaction]], and further injections were cancelled. Over the next 12&nbsp;days, Collip worked day and night to improve the ox-pancreas extract. A second dose was injected on 23 January, eliminating the [[glycosuria]] that was typical of diabetes without causing any obvious side-effects. The first American patient was [[Elizabeth Hughes Gossett|Elizabeth Hughes]], the daughter of U.S. Secretary of State [[Charles Evans Hughes]].<ref name=miracle>{{cite news | vauthors = Zuger A | title = Rediscovering the First Miracle Drug | url = https://www.nytimes.com/2010/10/05/health/05insulin.html | quote = Elizabeth Hughes was a cheerful, pretty little girl, five feet tall, with straight brown hair and a consuming interest in birds. On Allen's diet her weight fell to 65 pounds, then 52 pounds, and then, after an episode of diarrhea that almost killed her in the spring of 1922, 45 pounds. By then she had survived three years, far longer than expected. And then her mother heard the news: Insulin had finally been isolated in Canada. |work=[[The New York Times]] |date=4 October 2010 |access-date=6 October 2010 }}</ref><ref>{{cite web|url=https://insulin.library.utoronto.ca/islandora/object/insulin%3AM10011|title=Chart for Elizabeth Hughes| vauthors = Banting FG |date=16 August 1922|website=University of Toronto Libraries}}</ref>  The first patient treated in the U.S. was future woodcut artist [[James D. Havens]];<ref>{{cite web|url=https://insulin.library.utoronto.ca/islandora/object/insulin%3AT10060|title=Please save my son!| vauthors = Woodbury DO |date=February 1963|website=University of Toronto Libraries}}</ref> [[John Ralston Williams]] imported insulin from Toronto to [[Rochester, New York]], to treat Havens.<ref name="Marcotte">{{cite news| vauthors = Marcotte B |title=Rochester's John Williams a man of scientific talents |url=http://www.democratandchronicle.com/apps/pbcs.dll/article?AID=201011220301 |access-date=22 November 2010 |newspaper=[[Democrat and Chronicle]] |date=22 November 2010 |agency=[[Gannett Company]] |archive-url=https://archive.today/20101123001049/http://www.democratandchronicle.com/apps/pbcs.dll/article?AID=201011220301 |archive-date=23 November 2010 |location=[[Rochester, New York]] |pages=1B, 4B |url-status=dead }}</ref>

Banting and Best never worked well with Collip, regarding him as something of an interloper,{{citation needed|date=July 2021}} and Collip left the project soon after. Over the spring of 1922, Best managed to improve his techniques to the point where large quantities of insulin could be extracted on demand, but the preparation remained impure. The drug firm [[Eli Lilly and Company]] had offered assistance not long after the first publications in 1921, and they took Lilly up on the offer in April. In November, Lilly's head chemist, [[George B. Walden]] discovered [[Protein precipitation#Isoelectric precipitation|isoelectric precipitation]] and was able to produce large quantities of highly refined insulin. Shortly thereafter, insulin was offered for sale to the general public.

=== Patent ===
Toward the end of January 1922, tensions mounted between the four "co-discoverers" of insulin and Collip briefly threatened to separately [[patent]] his purification process. [[John G. FitzGerald]], director of the non-commercial public health institution [[Connaught Laboratories]], therefore stepped in as peacemaker. The resulting agreement of 25 January 1922 established two key conditions: 1) that the collaborators would sign a contract agreeing not to take out a patent with a commercial pharmaceutical firm during an initial working period with Connaught; and 2) that no changes in research policy would be allowed unless first discussed among FitzGerald and the four collaborators.<ref>{{cite web|url=https://insulin.library.utoronto.ca/islandora/object/insulin%3AW10033|title=Memorandum in reference to the co-operation of the Connaught Antitoxin Laboratories in the researches conducted by Dr. Banting, Mr. Best and Dr. Collip under the general direction of Professor J.J.R. Macleod to obtain an extract of pancreas having a specific effect on blood sugar concentration|last=University of Toronto Board of Governors Insulin Committee|date=25 January 1922|website=University of Toronto Libraries}}</ref> It helped contain disagreement and tied the research to Connaught's public mandate.

Initially, Macleod and Banting were particularly reluctant to patent their process for insulin on grounds of [[medical ethics]]. However, concerns remained that a private third-party would hijack and monopolize the research (as [[Eli Lilly and Company]] had hinted<ref>{{cite book | vauthors = Bliss M |title=The discovery of insulin |date=2007|publisher=University of Chicago Press|isbn=978-0-226-05899-3|edition= 25th anniversary |location=Chicago|pages=132|oclc=74987867|quote=The Lilly company would be delighted to work with Toronto, [[George Henry Alexander Clowes|Clowes]] wrote, and hinted, perhaps intentionally, perhaps not, that Toronto could be bypassed: "I have thus far refrained from starting work in our laboratories on the field of this question as I was anxious to avoid in any way intruding on the field of yourself and your associates until you had published your results. I feel, however, that the matter is now one of such immediate importance that we should take up the experimental end of the question without delay, preferably cooperating with you and your associates..."}}</ref>), and that safe distribution would be difficult to guarantee without capacity for quality control. To this end, [[Edward Calvin Kendall]] gave valuable advice. He had isolated [[thyroxin]] at the [[Mayo Clinic]] in 1914 and patented the process through an arrangement between himself, the brothers Mayo, and the [[University of Minnesota]], transferring the patent to the public university.<ref>{{cite web|url=https://insulin.library.utoronto.ca/islandora/object/insulin%3AL10262|title=Letter to Dr. J. J. R. Macleod 10/04/1922| vauthors = Kendall EC |date=10 April 1922|website=University of Toronto Libraries: Discovery and Early Development of Insulin}}</ref> On 12 April, Banting, Best, Collip, Macleod, and FitzGerald wrote jointly to the president of the [[University of Toronto]] to propose a similar arrangement with the aim of assigning a patent to the Board of Governors of the university.<ref>{{cite web|url=https://insulin.library.utoronto.ca/islandora/object/insulin%3AW10029|title=Statement read by J. J. R. Macleod at the Insulin Committee meeting regarding patents and royalties 28/04/1924| vauthors = Macleod JJ |date=28 April 1924|website=University of Toronto Libraries: The Discovery and Early Development of Insulin}}</ref> The letter emphasized that:<ref>{{cite book | vauthors = Bliss M |title=The discovery of insulin |date=2007|publisher=University of Chicago Press|isbn=978-0-226-05899-3|edition= 25th anniversary |location=Chicago|pages=131–133|oclc=74987867}}</ref>{{blockquote|The patent would not be used for any other purpose than to prevent the taking out of a patent by other persons. When the details of the method of preparation are published anyone would be free to prepare the extract, but no one could secure a profitable monopoly.}}The assignment to the University of Toronto Board of Governors was completed on 15 January 1923, for the token payment of $1.00.<ref>{{cite web |url= https://insulin.library.utoronto.ca/islandora/object/insulin%3AQ10013 |title=Assignment to the Governors of the University of Toronto| vauthors = Banting FG, Best C, Collip JS |date=15 January 1923|website=University of Toronto Libraries: Discovery and Early Development of Insulin}}</ref> The arrangement was congratulated in ''[[The World's Work]]'' in 1923 as "a step forward in medical ethics".<ref>{{cite web |url= https://insulin.library.utoronto.ca/islandora/object/insulin%3AW10031 |title=Copy of the article: A step forward in medical ethics|date=February 1923|website=University of Toronto Libraries: The Discovery and Early Development of Insulin|publisher=The World's Work}}</ref> It has also received much media attention in the 2010s regarding the issue of [[Health care prices in the United States|healthcare]] and [[Prescription drug prices in the United States|drug affordability]].

Following further concern regarding Eli Lilly's attempts to separately patent parts of the manufacturing process, Connaught's Assistant Director and Head of the Insulin Division [[Robert Defries]] established a patent pooling policy which would require producers to freely share any improvements to the manufacturing process without compromising affordability.<ref name="Bliss_2007">{{cite book |title=The discovery of insulin | vauthors = Bliss M |date=2007 |publisher=University of Chicago Press |isbn=978-0-226-05899-3 |edition= 25th anniversary |location=Chicago |pages=181 |oclc=74987867}}</ref>

=== Structural analysis and synthesis ===
{{multiple image
 | align = right
 | direction = vertical
 | width = 200
 | image1 = Insulin monomer 4INS.png
 | alt1 = Black-and-white ribbon diagram of a pig insulin monomer.
 | caption1 = [[Ribbon diagram|Richardson diagram]] of a [[pig|porcine]] insulin monomer, showing its characteristic [[secondary structure]]. This is the biologically active form of insulin.
 | image2 = Insulin hexamer 4INS.png
 | alt2 = Black-and-white ribbon diagram of a pig insulin hexamer, showing its characteristic quaternary structure. At the center is a pale blue-gray sphere representing a zinc atom.
 | caption2 = Richardson diagram of a porcine insulin hexamer. The sphere at the center is a stabilizing [[zinc]] atom, surrounded by coordinating [[histidine]] residues. This is the form in which insulin is stored in beta cells. {{PDB|4INS}}.
}}
Purified animal-sourced insulin was initially the only type of insulin available for experiments and diabetics. [[John Jacob Abel]] was the first to produce the crystallised form in 1926.<ref>{{cite journal | vauthors = Abel JJ | title = Crystalline Insulin | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 12 | issue = 2 | pages = 132–6 | date = February 1926 | pmid = 16587069 | pmc = 1084434 | doi = 10.1073/pnas.12.2.132 | bibcode = 1926PNAS...12..132A | doi-access = free }}</ref> Evidence of the protein nature was first given by [[Michael Somogyi]], [[Edward Adelbert Doisy|Edward A. Doisy]], and [[Philip Anderson Shaffer|Philip A. Shaffer]] in 1924.<ref>{{Cite journal| vauthors = Somogyi M, Doisy EA, Shaffer PA |date = May 1924 |title=On the Preparation of Insulin |url= https://www.jbc.org/content/60/1/31.full.pdf |journal=Journal of Biological Chemistry |volume=60 |issue=1 |pages=31–58 |doi = 10.1016/S0021-9258(18)85220-6 |doi-access=free }}</ref> It was fully proven when Hans Jensen and Earl A. Evans Jr. isolated the amino acids phenylalanine and proline in 1935.<ref>{{cite journal| vauthors = Jensen H, Evans EA |date=1 January 1935|title=Studies on Crystalline Insulin Xviii. the Nature of the Free Amino Groups in Insulin and the Isolation of Phenylalanine and Proline from Crystalline Insulin |url= https://www.jbc.org/content/108/1/1.full.pdf |journal=Journal of Biological Chemistry |volume=108 |issue=1 |pages=1–9 |doi=10.1016/S0021-9258(18)75301-5|doi-access=free }}</ref>

The amino acid structure of insulin was first characterized in 1951 by [[Frederick Sanger#Sequencing insulin|Frederick Sanger]],<ref name="Stretton_2002" /><ref name="sanger">{{cite journal | vauthors = Sanger F, Tuppy H | title = The amino-acid sequence in the phenylalanyl chain of insulin. I. The identification of lower peptides from partial hydrolysates | journal = The Biochemical Journal | volume = 49 | issue = 4 | pages = 463–81 | date = September 1951 | pmid = 14886310 | pmc = 1197535 | doi = 10.1042/bj0490463 }}; {{cite journal | vauthors = Sanger F, Tuppy H | title = The amino-acid sequence in the phenylalanyl chain of insulin. 2. The investigation of peptides from enzymic hydrolysates | journal = The Biochemical Journal | volume = 49 | issue = 4 | pages = 481–90 | date = September 1951 | pmid = 14886311 | pmc = 1197536 | doi = 10.1042/bj0490481 }}; {{cite journal | vauthors = Sanger F, Thompson EO | title = The amino-acid sequence in the glycyl chain of insulin. I. The identification of lower peptides from partial hydrolysates | journal = The Biochemical Journal | volume = 53 | issue = 3 | pages = 353–66 | date = February 1953 | pmid = 13032078 | pmc = 1198157 | doi = 10.1042/bj0530353 }}; {{cite journal | vauthors = Sanger F, Thompson EO | title = The amino-acid sequence in the glycyl chain of insulin. II. The investigation of peptides from enzymic hydrolysates | journal = The Biochemical Journal | volume = 53 | issue = 3 | pages = 366–74 | date = February 1953 | pmid = 13032079 | pmc = 1198158 | doi = 10.1042/bj0530366 }}</ref>  and the first synthetic insulin was produced simultaneously in the labs of [[Panayotis Katsoyannis]] at the [[University of Pittsburgh]] and [[Helmut Zahn]] at [[RWTH Aachen University]] in the mid-1960s.<ref name="Katsoyannis_1964">{{cite journal|vauthors=Katsoyannis PG, Fukuda K, Tometsko A, Suzuki K, Tilak M|year=1964|title=Insulin Peptides. X. The Synthesis of the B-Chain of Insulin and Its Combination with Natural or Synthetis A-Chin to Generate Insulin Activity|journal=Journal of the American Chemical Society|volume=86|issue=5|pages=930–32|doi=10.1021/ja01059a043}}</ref><ref name="pmid5881570">{{cite journal | vauthors = Kung YT, Du YC, Huang WT, Chen CC, Ke LT | title = Total synthesis of crystalline bovine insulin | journal = Scientia Sinica | volume = 14 | issue = 11 | pages = 1710–6 | date = November 1965 | pmid = 5881570 }} {{free access}}</ref><ref name="Marglin_1966">{{cite journal | vauthors = Marglin A, Merrifield RB | title = The synthesis of bovine insulin by the solid phase method | journal = Journal of the American Chemical Society | volume = 88 | issue = 21 | pages = 5051–2 | date = November 1966 | pmid = 5978833 | doi = 10.1021/ja00973a068 }}</ref><ref>{{cite journal | vauthors = Costin GE | title = What is the advantage of having melanin in parts of the central nervous system (e.g. substantia nigra)? | journal = IUBMB Life | volume = 56 | issue = 1 | pages = 47–9 | date = January 2004 | pmid = 14992380 | doi = 10.1080/15216540310001659029 | doi-access =free  | publisher = Time Inc. | s2cid = 85423381 }}</ref><ref name="isbn1-4020-0655-1">{{cite book |vauthors = Wollmer A, Dieken ML, Federwisch M, De Meyts P | title = Insulin & related proteins structure to function and pharmacology | publisher = Kluwer Academic Publishers | location = Boston | year = 2002 | isbn = 978-1-4020-0655-5 | url = https://books.google.com/books?id=Ula72_FSwy8C&q=Panayotis%20Katsoyannis&pg=PP11 }}</ref> [[Synthetic crystalline bovine insulin]] was achieved by Chinese researchers in 1965.<ref name="Zou2015">{{cite journal | vauthors = Tsou CL |author-link1=Chen-Lu Tsou |script-title=zh:对人工合成结晶牛胰岛素的回忆 |trans-title=Memory on the research of synthesizing bovine insulin|journal= 生命科学 [Chinese Bulletin of Life Science] |year=2015 |volume=27 |issue=6 |language=zh-hans |pages=777–79}}</ref> The complete 3-dimensional structure of insulin was determined by [[X-ray crystallography]] in [[Dorothy Hodgkin]]'s laboratory in 1969.<ref name=":2">{{cite journal | vauthors = Blundell TL, Cutfield JF, Cutfield SM, Dodson EJ, Dodson GG, Hodgkin DC, Mercola DA, Vijayan M | title = Atomic positions in rhombohedral 2-zinc insulin crystals | journal = Nature | volume = 231 | issue = 5304 | pages = 506–11 | date = June 1971 | pmid = 4932997 | doi = 10.1038/231506a0 | bibcode = 1971Natur.231..506B | s2cid = 4158731 }}</ref>

Hans E. Weber discovered preproinsulin while working as a research fellow at the University of California Los Angeles in 1974. In 1973–1974, Weber learned the techniques of how to isolate, purify, and translate messenger RNA. To further investigate insulin, he obtained pancreatic tissues from a slaughterhouse in Los Angeles and then later from animal stock at UCLA. He isolated and purified total messenger RNA from pancreatic islet cells which was then translated in oocytes from ''[[Xenopus laevis]]'' and precipitated using anti-insulin antibodies. When total translated protein was run on an SDS-polyacrylamide gel electrophoresis and sucrose gradient, peaks corresponding to insulin and proinsulin were isolated. However, to the surprise of Weber a third peak was isolated corresponding to a molecule larger than proinsulin. After reproducing the experiment several times, he consistently noted this large peak prior to proinsulin that he determined must be a larger precursor molecule upstream of proinsulin. In May 1975, at the American Diabetes Association meeting in New York, Weber gave an oral presentation of his work<ref>Weber, H.E. (1975) Diabetes 24, 405. (see figure)</ref> where he was the first to name this precursor molecule "preproinsulin". Following this oral presentation, Weber was invited to dinner to discuss his paper and findings by [[Donald Steiner]], a researcher who contributed to the characterization of proinsulin.  A year later in April 1976, this molecule was further characterized and sequenced by Steiner, referencing the work and discovery of Hans Weber.<ref>Chan SJ, Keim P, Steiner DF.  Cell-free synthesis of rat preproinsulins: Characterization and partial amino acid sequence determination.  Proc Natl Acad Sci. USA 1976;73:1964-1968.</ref> Preproinsulin became an important molecule to study the process of transcription and translation.

The first genetically engineered ([[Recombinant DNA|recombinant]]), synthetic human{{efn|Molecularly identical to human insulin: same sequence, same set of [[post-translational modifications]].}} insulin was produced using [[Escherichia coli|''E. coli'']] in 1978 by [[Arthur Riggs (geneticist)|Arthur Riggs]] and [[Keiichi Itakura]] at the [[Beckman Research Institute]] of the [[City of Hope National Medical Center|City of Hope]] in collaboration with [[Herbert Boyer]] at [[Genentech]].<ref name="urlGenentech" /><ref name="urlRecombinant DNA technology in the synthesis of human insulin" /> Genentech, founded by Swanson, Boyer and [[Eli Lilly and Company]], went on in 1982 to sell the first commercially available biosynthetic human insulin under the brand name [[Humulin]].<ref name="urlRecombinant DNA technology in the synthesis of human insulin"/> The vast majority of insulin used worldwide is biosynthetic recombinant human insulin or [[Insulin analog|its analogues]].<ref name="pmid23222785" /> Recently, another recombinant approach has been used by a pioneering group of Canadian researchers, using an easily grown [[safflower]] plant, for the production of much cheaper insulin.<ref>{{Cite web|url=https://www.ctvnews.ca/new-source-of-insulin-blossoming-on-the-prairies-1.479043|title=Safflowers may provide new insulin source {{!}} CTV News|website=www.ctvnews.ca|access-date=12 November 2019|date=February 2010}}</ref>

Recombinant insulin is produced either in yeast (usually ''[[baker's yeast|Saccharomyces cerevisiae]]'') or ''E. coli''. In yeast, insulin may be engineered as a single-chain protein with a [[Kexin|KexII endoprotease]] (a yeast homolog of PCI/PCII) site that separates the insulin A chain from a C-terminally truncated insulin B chain.  A chemically synthesized C-terminal tail containing the missing threonine is then grafted onto insulin by reverse proteolysis using the inexpensive protease trypsin;<ref name="pmid11030562">{{cite journal | vauthors = Kjeldsen T | title = Yeast secretory expression of insulin precursors | journal = Applied Microbiology and Biotechnology | volume = 54 | issue = 3 | pages = 277–86 | date = September 2000 | pmid = 11030562 | doi = 10.1007/s002530000402 | s2cid = 9246671 | url = http://w3.ualg.pt/~jvarela/biotecnol/pdf/humaninsulin.pdf | archive-url = https://web.archive.org/web/20170927000623/http://w3.ualg.pt/~jvarela/biotecnol/pdf/humaninsulin.pdf | archive-date = 27 September 2017 }}</ref> typically the lysine on the C-terminal tail is protected with a chemical protecting group to prevent proteolysis.  The ease of modular synthesis and the relative safety of modifications in that region accounts for common insulin analogs with C-terminal modifications (e.g. lispro, aspart, glulisine).  The Genentech synthesis and completely chemical synthesis such as that by [[Bruce Merrifield]] are not preferred because the efficiency of recombining the two insulin chains is low, primarily due to competition with the precipitation of insulin B chain.

=== Nobel Prizes ===
[[File:C. H. Best and F. G. Banting ca. 1924.png|thumb|left|[[Frederick Banting]] (right) joined by [[Charles Best (medical scientist)|Charles Best]] in 1924]]

The [[Nobel Prize]] committee in 1923 credited the practical extraction of insulin to a team at the [[University of Toronto]] and awarded the Nobel Prize to two men: [[Frederick Banting]] and [[John Macleod (physiologist)|John Macleod]].<ref name="urlThe Nobel Prize in Physiology or Medicine 1923">{{cite web | url = http://nobelprize.org/nobel_prizes/medicine/laureates/1923/ | title = The Nobel Prize in Physiology or Medicine 1923 | publisher = The Nobel Foundation }}</ref> They were awarded the [[Nobel Prize in Physiology or Medicine]] in 1923 for the discovery of insulin. Banting, incensed that Best was not mentioned,<ref>{{cite web | vauthors = Felman A | date = 22 November 2018 | title = Who discovered insulin? | work = Medical News Today | url = https://www.medicalnewstoday.com/articles/323774.php}}</ref> shared his prize with him, and Macleod immediately shared his with [[James Collip]]. The patent for insulin was sold to the [[University of Toronto]] for one dollar.

Two other Nobel Prizes have been awarded for work on insulin. British molecular biologist [[Frederick Sanger]], who determined the [[primary structure]] of insulin in 1955, was awarded the 1958 [[Nobel Prize in Chemistry]].<ref name="Stretton_2002">{{cite journal | vauthors = Stretton AO | title = The first sequence. Fred Sanger and insulin | journal = Genetics | volume = 162 | issue = 2 | pages = 527–32 | date = October 2002 | doi = 10.1093/genetics/162.2.527 | pmid = 12399368 | pmc = 1462286 }}</ref> [[Rosalyn Sussman Yalow]] received the 1977 Nobel Prize in Medicine for the development of the [[radioimmunoassay]] for insulin.

Several Nobel Prizes also have an indirect connection with insulin. [[George Minot]], co-recipient of the 1934 Nobel Prize for the development of the first effective treatment for [[pernicious anemia]], had [[diabetes]]. [[William Bosworth Castle|William Castle]] observed that the 1921 discovery of insulin, arriving in time to keep Minot alive, was therefore also responsible for the discovery of a cure for [[pernicious anemia]].<ref>{{cite journal | vauthors = Castle WB | title = The Gordon Wilson Lecture. A Century of Curiosity About Pernicious Anemia | journal = Transactions of the American Clinical and Climatological Association | volume = 73 | pages = 54–80 | year = 1962 | pmid = 21408623 | pmc = 2249021 | author-link = William Bosworth Castle }}</ref> [[Dorothy Hodgkin]] was awarded a Nobel Prize in Chemistry in 1964 for the development of [[crystallography]], the technique she used for deciphering the complete molecular structure of insulin in 1969.<ref name=":2" />

==== Controversy ====
[[File:Nicolae Paulescu - Foto03.jpg|thumb|upright=0.65|[[Nicolae Paulescu]]]]

The work published by Banting, Best, Collip and Macleod represented the preparation of purified insulin extract suitable for use on human patients.<ref name="pmid20314060">{{cite journal | vauthors = Banting FG, Best CH, Collip JB, Campbell WR, Fletcher AA | title = Pancreatic Extracts in the Treatment of Diabetes Mellitus | journal = Canadian Medical Association Journal | volume = 12 | issue = 3 | pages = 141–46 | date = March 1922 | pmid = 20314060 | pmc = 1524425 }}</ref> Although Paulescu discovered the principles of the treatment, his saline extract could not be used on humans; he was not mentioned in the 1923 Nobel Prize. Ian Murray was particularly active in working to correct "the historical wrong" against [[Nicolae Paulescu]]. Murray was a professor of physiology at the Anderson College of Medicine in [[Glasgow]], [[Scotland]], the head of the department of Metabolic Diseases at a leading Glasgow hospital, vice-president of the British Association of Diabetes, and a founding member of the [[International Diabetes Federation]]. Murray wrote:

{{blockquote|Insufficient recognition has been given to Paulescu, the distinguished [[Romania]]n scientist, who at the time when the Toronto team were commencing their research had already succeeded in extracting the antidiabetic hormone of the pancreas and proving its efficacy in reducing the hyperglycaemia in diabetic dogs.<ref name="pmid4560502">{{cite journal | vauthors = Drury MI | title = The golden jubile of insulin | journal = Journal of the Irish Medical Association | volume = 65 | issue = 14 | pages = 355–63 | date = July 1972 | pmid = 4560502 }}</ref>
}}

In a private communication, [[Arne Tiselius]], former head of the Nobel Institute, expressed his personal opinion that Paulescu was equally worthy of the award in 1923.<ref name="pmid4930788">{{cite journal | vauthors = Murray I | title = Paulesco and the isolation of insulin | journal = Journal of the History of Medicine and Allied Sciences | volume = 26 | issue = 2 | pages = 150–57 | date = April 1971 | pmid = 4930788 | doi = 10.1093/jhmas/XXVI.2.150 }}</ref>