An Interview with Manuel and Ann Varela: Edward Abraham and Antibiotic Synthesis

Mar 14, 2020 by

The Discoveries and Contributions of Edward Abraham—The Realm of Antibiotic Synthesis

Edward Abraham

Michael F. Shaughnessy –

1) Edward Abraham was born in England and studied in Southampton. When exactly was he born and where did he go to school?

Sir Dr. Edward Penley Abraham is primarily notable for his pioneering studies with the famous antibiotics penicillin and cephalosporin. Perhaps shockingly, Abraham is exceptionally well known for having discovered ways to destroy these medicines!

Abraham was born on the 10th of June, in 1913, in Shirley, Southampton, England to parents Maria Agnes and Albert Penley Abraham. While it is unclear where he attended grammar school, we know he attended King Edward VI High School. Abraham then attended Queen’s College, in Oxford, where he majored in chemistry, and in 1936, with honors, he took his degree in what amounted to the equivalent of both a college and a graduate master’s degree combined. Next, Abraham entered graduate school in Oxford’s Ph.D. program, where he focused his thesis project on the protein chemistry of lysozyme in the laboratory of his graduate advisor, Dr. Robert Robinson. Dr. Abraham took his doctorate from Oxford in 1938.

While in graduate school, Abraham met Asbjӧrg Harung, who was visiting Oxford from Norway. They married on the 1st of November in 1939 at Bergen, but the Second World War prevented them from being together. She managed to escape Nazi German occupation by foot through the mountains of Sweden in 1942. The couple’s marriage lasted until his death from a stroke on the 8th of May, 1999. Together, they had one child.

2) His name is almost synonymous with penicillin—Let’s set the record straight—who was the first to “discover “penicillin and what was Edward Abraham’s contributions.

While Abraham’s contributions to the study of penicillin are numerous, he was certainly not the first investigator to discover the antibiotic. The credit for penicillin’s discovery goes to Dr. Alexander Fleming, whom you’ll recall was featured in chapter 9 of our book “The Inventions and Discoveries of the World’s Most Famous Scientists.” As you know, in 1928, Fleming serendipitously discovered that the juice from a contaminating mold on his Petri dish killed Staphylococcus bacteria. One of the most historically essential antibiotics, penicillin, had been uncovered. For his innovative discovery, Fleming took the Nobel Prize in 1945.

Abraham’s contributions to this field of penicillin studies are several-fold. First, he was a junior member of the team that was dedicated, in 1940, to the purification of penicillin. He worked as an assistant to Drs. Howard Florey and Ernst Chain during their antibiotic purification work. The purified penicillin was then used in clinical studies on humans to test its safety.

Later studies addressed the efficacy of penicillin towards bacterial infection—see below. Second, Abraham collaborated with Chain in 1940, and together they discovered a substance produced by crushed cells of the bacterium Escherichia coli (better known at the time as Bacillus coli), which destroyed the penicillin. Abraham and Chain deduced that the nature of this destructive power was proteinaceous.

They further speculated that the protein was an enzyme that metabolized the penicillin, converting it into a useless derivative. Thus, they named this enzyme penicillinase. Later studies showed that penicillinase belonged to a family of enzymes known as β-lactamases.

A third noteworthy contribution by Abraham involves a clinical study in 1941 in which their purified penicillin was tested in human patients for efficacy in the treatment of bacterial infection. The new penicillin treatment significantly improved the patients’ clinical conditions! It was a pioneering study.

Lastly, Abraham’s participation in the penicillin project led to the determination of its molecular structure. Interestingly, in the center of the structure, the “square” lies its active site.

Structures of penicillin.

The active site contains a 4-membered ring consisting of three carbons and one nitrogen atom, forming a so-called β-lactam ring. For penicillin to actively kill certain its target bacteria, the center β-lactam ring structure must be intact; otherwise, the antibiotic will not work against specific bacterial infections.

Abraham’s penicillin structure was met with disbelief. During the time that the famous penicillin structure was determined by Abraham and colleagues, its scientific accuracy had come into considerable question by Robinson, his graduate advisor who was considered a genius and seldom, if ever, mistaken. In fact, Robinson proposed his own version of the penicillin structure but later shown to be erroneous.

In 1944, Dr. Robert Burns Woodward conducted chemically-based experiments that supported the structural nature of the β-lactam ring. In 1949, Dr. Dorothy Crowfoot Hodgkin confirmed the precise molecular configuration of penicillin using X-ray crystallographic analysis.

3) Moving along—he apparently was also involved with cephalosporin- what was his contribution in this realm?

In 1945, Dr. Giuseppe Brotzu discovered the antibiotic cephalosporin. He isolated a mold called Cephalosporium acremonium that lived in a nearly pristine body of seawater near a sewage outlet located along the coast of Cagliari, Sardinia. The contemporary name for this fungus is Acremonium chrysogenum. In 1948, Brotzu found that the fungus secreted the antibiotic and that it inhibited the growth of both Gram-negative and Gram-positive bacteria. He was, however, unable to expand his efforts to develop the new antibiotic. This is where Abraham steps into the picture. Brotzu sent his fungus to the laboratory where Abraham was working.

Working with Guy G.F. Newton in the laboratory of Professor Sir Howard Florey at Oxford, Abraham participated in studies devoted to the development of the new antibiotic. In 1949, they grew large batches of Brotzu’s fungus and purified several compounds, which they named cephalosporins P1, P2, P3, P4, and P5. They used the designation P because the agents could kill Gram-positive bacteria. These findings were published in 1951. One problem, however, was that none of these antibiotics killed Gram-negative bacteria, as Brotzu had initially reported. Thus, their work on cephalosporin development continued.

In 1950, they purified an additional substance from Brotzu’s fungus. They called the new substance cephalosporin N because it killed Gram-negative bacteria. When this new anti-bacterial agent was analyzed structurally, it was found to be another penicillin. So they renamed it penicillin N and published the work in 1954. During Abraham’s work with penicillin N purification, however, they noticed an emission peak in their spectral analysis, indicating another agent. Thus, they purified it, too, and found that their new cephalosporin C could kill Escherichia coli, Salmonella enterica Typhi, and Staphylococcus aureus.

Structure of cephalosporin C

Similar to their encounter with the disbelief about their penicillin structure, the nature of cephalosporin C’s structure determined by Abraham and Newton, published in 1961, likewise proved to be just as controversial. This time, doubts about the true nature of the new antibiotic’s structure had emerged from a prominent faculty member of Harvard’s chemistry department, Dr. Robert Burns Woodward. Like Robinson before him, Woodward was widely considered by organic chemists as another genius. Previously, Woodward had also solved the correct structure of penicillin, as had Abraham. This time, however, Woodward immediately criticized Abraham’s cephalosporin C structure.

Nevertheless, an undeniable experimental confirmation of the Abraham-Newton structure arose from Drs. Dorothy Hodgkin and Rex Richards, who used X-ray crystallography. They definitively demonstrated that cephalosporin C had the β-lactam ring in its center, and as Newton and Abraham originally predicted, all remaining chemical constituents. Therefore, Abraham had managed to correctly deduce the structures for two critically important antibiotics, penicillin and cephalosporin C, even while other arguably brilliant scientists, such as Robinson and Woodward, had been wrong.

Moving on, Abraham used cephalosporin C as a platform upon which to make new anti-bacterial derivatives chemically. The goal was to make the bacteria-killing power of cephalosporin C more efficient by chemical modification. One of these modified spin-offs was the synthetic compound called 7-ACA, which was short for 7-aminocephalosporanic acid. The new spin-off molecule was classified as a semi-synthetic, because a naturally occurring agent, the antibiotic cephalosporin C, was changed by artificial chemical means (synthetic) in the laboratory.

The new 7-ACA proved to be worth the effort because it was quite effective in killing bacteria! Abraham’s discovery turned out to be the first semi-synthetic anti-bacterial medicine of the cephalosporin class that became commercially available. Some sources maintain the assertion that Abraham’s chemical construction of 7-ACA was actually the first semi-synthetic ever produced.

In any case, his new medicine was later to be called cephalothin, and it was a model member of the first generation of cephalosporins. New generations of cephalosporins have since been developed—as of the latest count, five such generations exist. As a whole, the cephalosporins are among the most widely prescribed β-lactam antibiotics in modern medicine. In recent times, Abraham’s cephalothin is still covered in medical and biochemistry textbooks.

4) Apparently, cephalosporin can treat penicillin-resistant bacteria. First, why would some bacteria become resistant, and what does cephalosporin do differently, or how does it address penicillin-resistant bacteria?

Bacteria develop and acquire resistance to antibiotics for at least two established reasons.

To begin with, certain bacteria themselves produce antibiotics, and to live and grow in the presence of the poisons they make, they will need to avoid their toxic effects. Thus, antibiotic resistance mechanisms permit the survival even in the presence of sub-growth inhibitory or seemingly lethal amounts of their home-grown antibiotics.

Second, the production and release of homemade antibiotics by specific bacteria permits them to stop the growth or even kill neighboring microbes, bacteria of which could most certainly compete for any limited resources, such as food, water, ions, or certain growth factors. Thus, bacteria harbor antibiotic resistance to beat out their nearby competitors for survival.

Penicillin is known to kill bacteria by encouraging them to explode into pieces. The famous antibiotic binds to and prevents the so-called penicillin-binding proteins (PBPs), also known as transpeptidases, in the cell wall of bacteria and prevent them from making strong cell walls. These bacteria with weak cell walls permit their explosions as they cannot withstand the internal osmotic pressure produced by their growth and metabolic activities. Thus, penicillin-susceptible bacteria undergo lysis and die.

One prime reason for penicillin resistance in bacteria is that they produce penicillinase enzymes. These enzymes, also known as β-lactamases, metabolically destroy active penicillin into inactive penicilloic acid. This damaged form of the drug cannot kill bacteria. Another reason for bacterial penicillin resistance is that the main target of the drug, the transpeptidases (or PBPs), changes its shape. That is, the penicillin’s cellular target has been altered, and it can no longer bind to the PBPs (or binds weakly), permitting the resistant bacterium to live and grow with its strong cell wall.

The molecular natures of the bacterial cellular target for penicillins and cephalosporins are the same: the PBPs or transglycosylases. Nevertheless, as you pointed out, specific cephalosporins can still kill certain penicillin-resistant bacteria. A number of causes have been invoked to explain this phenomenon. One reason is that the newer cephalosporins were better able to enter the cells of penicillin-resistant bacteria, allowing them to bind their target and lyse the bacteria.

Another possibility is that the cephalosporins could attach to the targets more efficiently and stay bound to the PBPs. The tighter binding to the transpeptidases rendered them ineffective in producing strong cell walls, and the bacteria then underwent lysis. A third reason could be that the penicillinases did not work as well on the cephalosporins, leaving them somewhat alone and thus allowing them to proceed onto their targets to lyse the bacteria.

5) Pneumonia, bronchitis, and septicemia are the main conditions that cephalosporin is used to treat. What is it about these conditions, and how does cephalosporin work exactly?

Pneumonia is an inflammatory response in the lungs to bacterial infection. Patients with pneumonia can experience a build-up of pus and other fluids in the lung, making it challenging to breathe adequately. The condition can be dangerous in individuals who are in a weakened state, such as those who are elderly or very young. It can also be severe in patients who are immunocompromised or immunodeficient.

Likewise, bronchitis is an inflammation of the bronchioles, which are the tubes that permit breathing of air into and out of the two lungs. Bronchitis can acute (and short-lived) or chronic (and more dangerous).

Septicemia is a condition where bacteria are present and actively growing in a patient’s blood. If bacteria are merely present in the blood but not increasing their numbers, the condition is called bacteremia. Sepsis refers to a patient’s response to septicemia and may involve the immune response. Both septicemia and sepsis are dangerous situations for patients who experience them.

The cephalosporin antibiotics and their semi-synthetic derivatives have enjoyed a certain level of success in treating these and other clinical conditions caused by bacterial pathogens. The prime cellular target in bacteria of the cephalosporins is the microbial enzyme called transpeptidase.

Bacteria harbor these transpeptidase enzymes in order to build a critical component of their cell walls, called peptidoglycan, in order to ensure their maximum protection. The transpeptidase enzyme catalyzes the crosslinking of two peptidoglycan parts, called N-muramic acid (NAM), to each other. When the two NAMs are crosslinked, pentapeptides are formed between them. These pentapeptides consists of five amino acids. The NAM cross-linked parts form deep layers to build the protective peptidoglycan of the bacterial cell wall. Cephalosporin prevents this pentapeptide crosslinking, thus preventing the bacterium from constructing a strong protective cell wall. Such bacteria with their weak cell walls have a propensity to explode.

While I had mentioned above that some penicillin-resistant bacteria are susceptible to certain cephalosporins, there are indeed specific β-lactamases that do indeed metabolize them to inactive states as well. Such circumvention of the lethal nature of the cephalosporins can allow resistant bacteria to grow and wreak havoc in patients infected with them.

Some bacteria have actually developed extended-spectrum β-lactamases (ESBLs), which digest many types of β-lactams and at high rates of degradation. Bacteria that harbor ESBLs can be quite bothersome to some patients, if not lethal.

6) There apparently have been about five revisions or five generations of cephalosporin that have been shown effective against MRSA. First, what exactly is MRSA, and can you discuss what scientists mean by these “second generation” drugs. I take it that each revision improves the drug.

The term MRSA signifies methicillin-resistant Staphylococcus aureus. The MRSA bacteria have emerged as severe pathogens in clinical settings, causing healthcare-associated infections (HAIs). These HAIs are particularly problematic because they may occur in patients who enter a hospital for whatever medical reason but acquire an MRSA infection in the meantime. The methicillin used to be a clinically useful antimicrobial agent, until the MRSA emerged, that is, in which the antibiotic no longer effectively treated Staphylococcus aureus infections.

Exacerbating the problem of MRSA bacteria, these microbes can also be resistant to many other classes of antibiotics, making them potentially untreatable in their patients. To intensify this issue, MRSA moved out of the clinical settings and established themselves in the general community. Such MRSA pathogens are also referred to as community-acquired Staphylococcus aureus (CASA).

At the time of the development of the cephalosporins, these first-generation antibiotics, e.g., Abraham’s cephalothin and cephazolin, were warmly welcomed and used in the clinical setting to treat bacterial infections effectively. However, almost immediately after their implementation, bacterial resistance to these first-generation cephalosporins were reported in the medical literature.

Thus, a second-generation of the cephalosporins, namely, cefuroxime and cefoxitin, were devised. As with the first generation, these new drugs were used in the clinical arenas and shown to be initially useful during the treatment of infectious disease. However, soon after their implementation, bacterial resistance emerged with cefuroxime and cefoxitin, just as had been reported with the first-generation agents.

Therefore, the third generation of cephalosporins was developed, synthesizing cefotaxime ceftriaxone, and ceftazidime. Again, the same story happened with these new drugs as had occurred with the previous two generations of medicines. Likewise, the fourth (cefepime and cefpirome) and fifth-generation (ceftobiprole and ceftaroline) cephalosporins experienced the same sort of situation as before. The cycle of semi-synthesis, clinical usage, treatment efficacy, and bacterial resistance seems to be never-ending.

7) Apparently, in addition to being a scientist- he was also a philanthropist- What can you tell us about his generosity?

Indeed, garnering of patents and commercialization of the cephalosporins could have made Abraham extremely wealthy. In published tributes to Abraham, colleagues have described him as modest, reserved, kindhearted, gentle, and even self-deprecating. Rather than pocketing an enormous personal wealth, he paved a path toward philanthropy.

He worked to establish several charitable funding avenues. One such channel was the Edward Abraham Fund, which was geared towards helping medical chemistry and biology studies at several research institutions at Oxford. Another road was the EPA Cephalosporin Trust, which was targeted to the educational plus research areas at Oxford, King Edward’s School, his alma mater, as well as the Royal Society.

Abraham collaborated with Newton to inaugurate the Guy Newton Research Fund in 1970. The foundation was for the benefit of research studies at the Sir William Dunn School of Pathology.

These and other charitable organizations inspired by Abraham are credited to have benefited many higher educational institutions notable to him as well as the overall field of medical biochemistry. Abraham has undeniably made a substantial philanthropic contribution in clinical medical research and has left a long-lasting legacy because of it.

While Abraham chose not to benefit personally in a monetary fashion by his antibiotic work, he was, nonetheless, awarded in other ways. He was inducted into the prestigious Royal Society in 1958, and in 1973 this organization bestowed upon Abraham the coveted Royal Medal. In 1980, the Society honored Abraham with the Mullard Award, and in 1983, he became an honorary fellow as part of the American Academy of Arts and Sciences.

8) His legacy is stunning. In your mind, what were his major contributions to the realm of biochemistry?

Sir Edward P. Abraham will forever be known because of his pioneering work on the penicillin and cephalosporin antibiotics. His scientific legacy also entails the first discovery of early clinically useful antibiotic derivatives. New generations of scientists are working on new generations of antibiotics, taking the mantle started by Abraham and continuing the work. These antibiotics and their semi-synthetic relatives are still of tremendous clinical utility in modern times. Because of Abraham, countless millions of lives have been spared from the lethal effects of bacterial pathogens for several generations.

Following the lead inspired by Abraham, biochemists will be needed to continue the work of finding new microbes with novel antibiotics. Biochemists will then be required to purify these new putative antimicrobial agents and to determine their molecular structures, just as Abraham did. Further, biochemists will need to study the chemistry of these novel antibiotics to learn how they function in killing or inhibiting microbes. Then, just as Abraham had established, chemists will need to modify individual sites on the antimicrobial molecules in order to synthesize new derivatives with better anti-bacterial activities, in order to treat infections better.

Along the way, Abraham also helped to discover various means that bacteria use to develop resistance to each of these agents. It was necessary to understand these antibiotic resistance mechanisms in order to find ways to circumvent them.

This research paradigm, pioneered by Abraham, will no doubt be of relevance for many generations of biochemists and medicines, alike.

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