Chris Anthony

J Rodney Quayle was an outstanding microbial biochemist whose early training in pure chemistry was coupled with rigorous enzymology and experience in the relatively new techniques of using radioactive 14C compounds in the study of metabolic pathways. These he used to investigate and elucidate the pathways of carbon assimilation during microbial growth on compounds with a single carbon atom such as methane and methanol. When he started, little was known about these methylotrophs which, largely as a result of his own work and the work inspired by him, have formed the subject of regular international symposia over a period of more than 40 years. After a short time working in Melvin Calvin’s lab in California and a very fruitful period in Krebs’s Unit for Research in Cell Metabolism in the University of Oxford he moved for the next twenty years to the University of Sheffield, after which he became a highly successful and popular Vice-Chancellor at the University of Bath. His rigorous approach to his subject, his generosity and inspiration made him a much revered and much loved father figure to generations of microbial biochemists.

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                                                          EARLY LIFE

John Rodney Quayle (always known as ‘Rod’) was born on November 18, 1926, in Hoylake, a seaside town located at the north western corner of the Wirral Peninsula on Merseyside, England. He and his brother, Brian, lived with the family over his father’s pharmacist shop. Brian was only 14 months older than Rod and they were very close. Brian also had a successful career, becoming an executive for IPC Magazines in London.
Rod’s maternal grandfather was also a pharmacist with his own shop nearby in Hoylake. Rod has said that maybe his early exposure to a world of slender flasks filled with variously coloured liquids, measuring cylinders and chemical balances led to his later interest in chemistry.
       For reasons that he never really understood, his parents separated in 1931 and he, Brian and his mother moved in with his recently retired maternal grandparents to their holiday bungalow in Cilcain, a small farming village on the side of the Clwydian range of hills in a very rural setting about four miles from the market town of Mold in Flintshire, North Wales. Rod never did find out the reason why his father had left, or where he had gone. His father did not support the family financially, and his grandfather had to support them all out of the money he had saved for retirement, hence money was always very short. Rod said that, although their lack of a father led to teasing at school, Cilcain was a wonderful place to live, wandering about the countryside with none of the constraints that children are placed under today.
       They attended the village (Church of England) primary school with about 30 pupils, mostly farmers’ children. They were taught by three spinster schoolmistresses; all the infant classes were taken by one teacher in one room, and all of the other children were taught by the other two teachers in another room at the same time. At the age of 10 Rod sat the entrance examination for the Alun Grammar School in Mold, achieving third place in an intake of about 50 entrants. Many of his schoolfellows were wartime evacuees from Merseyside.
       In 1943 he gained a Higher School Certificate of the Central Welsh Board with a distinction in Physics, a credit In Chemistry and a pass in Mathematics, leading to the award of a County Scholarship for University study. That summer he was not yet 17 and his headmaster and his admired and influential physics master (G. Roblin) advised him to go straight up to University and at least complete the first year before possible military service took over. So in September of that year he entered University College of North Wales at Bangor, a coastal city between the Menai Strait and the mountainous Snowdonia National Park, providing an environment that fostered his love of mountain walking and rock climbing. He also become a member of the Bangor rowing team, an activity he continued when he moved to Cambridge where he took it very seriously, in the ‘Gentlemen’s Rowing Team’.
       After graduating with a B.Sc. (Honours) in Chemistry in 1946, Rod stayed on at Bangor to take a Ph.D. in Physical Organic Chemistry. During this time he was awarded a University of Wales Post-graduate Studentship and was appointed Tutor at the Hall of Residence (Neuadd Reichel). His research was supervised by Professor E.D. Hughes (FRS 1949), a trailblazer in kinetics and mechanisms in organic chemistry; the title of Rod’s thesis was “The reactions of s‑triphenylbenzene and derivatives with special reference to steric hindrance”. Based on this research he was awarded a University of Wales Fellowship which persuaded Professor Alexander (later Lord) Todd (FRS 1942 PRS 1975-1980) to accept him into his laboratory at the University of Cambridge to work on the structure and chemical synthesis of the blood pigments of Aphids (Aphididae) which led to ten publications, mainly in the Journal of the Chemical Society. While there he entered St. John’s College as a research student leading to the award of a second PhD degree (in 1952) with a thesis entitled “Colouring Matters of the Aphididae".  In 1951 he was awarded a Senior Research Award from the Department of Scientific and Industrial Research (DSIR) and in the same year he married Yvonne Sanderson, who had been a fellow student at Bangor.

INTRODUCTION TO METHYLOTROPHY

After Cambridge, Rod’s research changed direction towards enzymology and microbial biochemistry. This paragraph has therefore been inserted in order to place his subsequent contributions into this context. Any account of his contribution to microbiology becomes a broad chronological survey of those bacteria and yeasts that grow at the expense of compounds containing a single carbon atom (C1 compounds) including methane, methanol and methylamine. When he started this research, very few such micro-organisms had been described but it is now known that there exists a huge diversity of these microbes having a correspondingly wide metabolic diversity. All living organisms are either autotrophs or heterotrophs. Autotrophs use carbon dioxide as their sole source of carbon for growth while heterotrophs use organic carbon sources. The name methylotrophs, was coined to include the huge group of heterotrophs that are able to grow at the expense of reduced C1 compounds. The groups of microbiologists with an interest in these previously unknown bacteria has expanded enormously over the years, so at the time of writing there have been 17 international symposia and conferences on this topic. For reviews of his contribution to the study of methylotrophy see his reviews (1, 2, 3, 4) and for a more extensive account see Anthony (1982). 
       An admirable characteristic of Rod’s research presentations was his concern to emphasise the importance of the contributions of his students and associates. Much of the work described here is based on these contributions and to achieve a clearer presentation his colleagues are not always mentioned but can be identified in the complete list of publications.

CALIFORNIA

In 1953, Rod was awarded a Fulbright Travel Grant, to work as a Research Fellow with Melvin Calvin (Foreign Member FRS 1959) in the Department of Chemistry of the University of California at Berkeley, concentrating on the first enzyme-catalysed step in the assimilation of carbon dioxide by plants. It was here that he established a friendship with Hans Kornberg that was to have important relevance later in his career.
Although he was not to know at the time, Rod’s path to methylotrophy can be said to have started in the Calvin laboratory in Berkeley (1953-1955). It was here that the Calvin-Benson Cycle was established as the photosynthetic pathway for assimilation of carbon dioxide in plants. Rod became the first author of a publication (5) still regarded as providing conclusive evidence for the existence of this cycle. It had been shown that plants fixed 14C-labeled carbon dioxide initially by carboxylation of ribulose 1,5-bisphosphate (RuBP) (first synthesized by Rod) to give an intermediary C6 compound that was cleaved into two C3 compounds. He showed that cell-free extracts of the green alga Chlorella , exposed to labelled CO2 for only 1 minute, formed labelled phosphoglycerate when RuBP was also added, and that phosphoglycerate was not formed when alternative candidate sugar phosphates were used. The authors of this one-page communication (5)concluded, "It is clear that the extracts contain an enzyme (or enzymes) capable of catalyzing the carboxylation of ribulose diphosphate, specifically, to form phosphoglyceric acid”. This was the first description of the activity of the enzyme now known as ribulose bisphosphate carboxylase (RuBisCo), the most abundant enzyme on Earth. It was in Berkeley that Rod first met Hans Kornberg (later Sir Hans Kornberg FRS 1965) with whom he would later collaborate in Oxford. Rod impressed Calvin not only by his excellent science but, as mentioned in a reference posted to Hans Krebs (later Sir Hans Krebs FRS 1947) to support his application to join the MRC Unit in Oxford, “His stabilizing influence was felt throughout the lab, particularly by the graduate students as well as by his peers”.

OXFORD

In 1955, Rod and Yvonne returned to England where Rod had accepted a post as Senior Scientific Officer in the DSIR's Tropical Products Institute in London, to study the chemistry of the naturally occurring pyrethrin insecticides. He was not enthusiastic about the topic or the rigid constraints of work in a government laboratory, and within a year a chance meeting with Hans Kornberg at a London theatre led him to apply for a position with Professor Hans Krebs in his MRC Unit for Research in Cell Metabolism at the University of Oxford. Here he enjoyed a fruitful collaboration with Kornberg on the newly-proposed glyoxylate cycle for growth of bacteria on C2 compounds. This led to what became his main research area - the biochemistry and physiology of bacteria growing on the C1 compounds: formate, methane, methanol and methylamine.  Although a member of a Research Unit within the precincts of Oxford University he was not part of it until 1957 when he accepted a Lectureship in Biochemistry at Oriel College. With it came formal membership of the University, offering an opportunity to demonstrate his exceptional gifts as an inspiring and caring teacher and research supervisor.

Bacterial growth on C2 compounds

Working in Krebs’s MRC Unit, Hans Kornberg had addressed the question of how bacteria are able to grow on the C2 compound acetate. The tricarboxylic acid (TCA) cycle (also known as the Krebs Cycle) functions to oxidise acetate to CO2 but acetate also has to be converted into C3 and C4 compounds for assimilation into cell material. To achieve this, supplementary enzymes are needed to replenish the cycle when intermediates are removed for biosynthesis. The glyoxylate cycle (or ‘bypass’) had recently been proposed by Kornberg and Krebs (1957), comprising two key anaplerotic (‘filling up’) enzymes - isocitrate lyase and malate synthase, previously described in the literature, and they showed both to be present in cell-free extracts of acetate-grown bacteria. It was still necessary to demonstrate that the cycle operates in vivo.  Using the chromatographic techniques that had proved so successful in Berkeley for elucidating the Calvin-Benson Cycle in photosynthesis, Rod and Hans Kornberg joined forces to isolate and characterise by chemical degradation, intermediates formed from 14C-labelled acetate and CO2. The isotope distributions supported the simultaneous operation of the TCA and glyoxylate cycles (6). Such experiments became the key type of experiment subsequently used by Rod in his future work on methylotroph pathways.
       After their initial collaboration they made independent studies on the metabolism of C2 compounds more highly oxidized than acetate.  Rod was joined by an Australian visitor, Bruce Keech, to investigate the routes by which Pseudomonas oxalaticus grows on oxalate, the most oxidized C2 compound. In a series of seven papers he described how oxalate was reduced to glyoxylate and then converted into glycerate (the precursor of other cell constituents) by the glycerate pathway. The glycerate pathway had been discovered by Kornberg and his student Tony Gotto working on the other side of their shared laboratory bench using glycollate-grown Pseudomonas ovalis; two of their key papers on bacterial growth on glycollate (Kornberg) and oxalate (Quayle) were published together in the same issue of Nature in 1959.  Rod’s last paper in this series showed that energy for growth comes from decarboxylation of oxalyl-coenzyme A to formate and thence to carbon dioxide. A key intermediate in this process is a tiny formyl group bonded to two large coenzymes, coenzyme A and thiamine pyrophosphate. A visitor to his lab at that time was met by Rod bubbling with excitement because of this intermediate “Which showed so much care being taken over one little carbon atom”.

Bacterial growth on C1 compounds

Pseudomonas oxalaticus is also capable to growth on formate as its sole source of carbon and energy, raising the question of how this C1 compound is assimilated into cell material. The obvious route would be by way of the Calvin-Benson photosynthetic pathway. Again with Bruce Keech, Rod showed that this is indeed the case. The first intermediate in the assimilation of 14C‑formate or 14C‑bicarbonate was phosphoglycerate, the formate being oxidised to carbon dioxide before assimilation into cell material. Similar extensive experimentation, as he had performed in Calvin’s lab using Chlorella extracts, confirmed the presence of the key enzyme RuBP carboxylase together with phosphoribulokinase during growth of P. oxalaticus on formate but not on oxalate.
        About fifteen years later he showed that some methylotrophs (e.g. Paracoccus denitrificans) also grow on methanol by way of the ribulose bisphosphate pathway (the Calvin-Benson photosynthetic pathway).
       Rod has pointed out that his study of methylotrophic microbes might have taken a different route had he started with Paracoccus instead of isolating his own organism, Pseudomonas AM1 (now Methylobacterium extorquens AM1). This organism was originally isolated on a methylamine medium in 1960 by his first research student, David Peel.  After 50 years it is still the ‘workhorse’ of those working on the biochemistry, molecular biology and biotechnology of bacteria growing on methanol. He used the same experimental approaches that he had developed to confirm the operation of both the glyoxylate cycle during growth on acetate and the glycerate pathway for assimilation of oxalate. With his students Peter Large and David Peel, in a series of elegant and rigorous studies (see 7), he showed that methanol is assimilated in Pseudomonas AM1 (and in Hyphomicrobium vulgare) by a novel assimilatory pathway, the serine cycle, in which methanol is fixed at the oxidation level of formaldehyde into serine, and at the level of carbon dioxide into oxaloacetate. This model investigation involved the identification of early intermediates produced by incubating methanol-grown bacteria with either 14C-methanol or 14C-bicarbonate, the position of the isotope in the isolated labeled compounds (serine, glycine, aspartate etc) being determined by chemical degradation and analysis (Figures 1 and 2). The pathway was then established by the discovery and detailed characterization of novel specific enzymes characteristic of the pathway, their importance being confirmed by studying their induction during methylotrophic growth and by their absence in mutants lacking the ability to grow on methanol. By the mid 1970s the serine cycle appeared to be complete, except for understanding how acetyl-coenzyme A is oxidised to glyoxylate.  This essential step was not elucidated for another 30 years.  After each International Conference on Microbial Growth on C1 compounds in the intervening period, Rod would ask “Have they solved the serine cycle yet?” Sadly it was not until 2006, shortly after Rod had died, that a satisfactory description of the complete pathway arose from a presentation by George Fuchs and the Freiburg group at the Gordon Research Conference in Magdalen College, Oxford. For a description of the development and history of Rod’s serine cycle see his influential 1972 review (2) and for a more extensive review of this cycle and its completion see Anthony (2011).

Figure 1. Figures taken from the first paper published by Rod Quayle (with P. J. Large and D. Peel) on methanol assimilation by Methylobacterium extorquens AM1 (current name) (4). They show the result of experiments in which samples were taken every few seconds after the addition of radioactive substrate, after which the labelled metabolic intermediates were separated, identified and quantified. A key feature of these experiments is that it is not the total activity in each intermediate that is graphically recorded but the percentage of label in each compound. This
decreases with time for an early intermediate in the pathway. (Reproduced courtesy of Biochemical Journal)

Figure 2. Figure taken from the second paper published by Rod Quayle (with P. J. Large and D. Peel) on methanol assimilation by Methylobacterium extorquens AM1 (current name) (5). This shows an outline of the two possible pathways for methanol (and carbon dioxide) assimilation, based on the experiments illustrated in figure 1 together with analysis of the locations of the radioactive carbon atoms within the postulated intermediates. Rod eventually showed that the assimilation pathway is cyclic (the serine cycle). (Reproduced courtesy of Biochemical Journal)

MICROBIOLOGY IN SHEFFIELD (1963 – 1983)

Professor Hans Krebs FRS was the head of the Biochemistry Department in the University of Sheffield (1938 – 1954) where he frequently used microorganisms in his research, appreciating their many advantages for studying cell metabolism. In 1948/9 he helped establish within the Bacteriology Department, a Sub-Department of Microbiology with Sidney Elsden as the Senior Lecturer-in-charge and one lecturer (Bernard Fry). The Department of Microbiology was then formally established in 1952, and the Agricultural Research Council added a Unit of Microbiology with Elsden as its Honorary Director and three additional members of ARC staff. Krebs was later involved in establishing the West Riding Chair of Microbiology for Sidney Elsden (1959-65), the title reflecting generous financial support from the West Riding County Council. During its first 13 years (1952-1965) the Department of Microbiology gained international recognition for research on the biochemistry of anaerobic and photosynthetic bacteria and the energetics of bacterial growth and it ran a successful M.Sc. course in Microbiology.
       In 1954 Krebs moved with most of his staff from Sheffield, to take the Whitley Chair of Biochemistry at Oxford. His replacement, Professor Quentin Gibson, with an extremely successful group of scientists quickly re-established the Department’s prestigious international reputation.  Unfortunately the Sheffield Vice-Chancellor's failure to value their exceptional quality coupled to attractive funding opportunities in North America, led to a seemingly infamous exodus in 1962/3. The Sheffield Biochemistry Chair was then filled by Walter Bartley (1963-81) and, as the future of the Oxford MRC Unit became uncertain due to Krebs’s impending retirement, Rod accepted an invitation to move with Bartley to the Sheffield Department as a Senior Lecturer.
       Two years later Rod was appointed to the West Riding Chair of Microbiology and Head of the Microbiology Department (1965-83) following Elsden’s move to Norwich as Director of the ARC Food Research Institute with the simultaneous departure of his ARC Staff.  Three Lecturers (John Guest, Margaret Attwood and Peter White) were soon appointed to replace the former ARC Staff and with Bernard Fry as Senior Lecturer they represented the entire research and teaching staff of the Department for the next ten years. They were joined later by Milton Wainwright (1975) and Ann Moir (1981) ultimately bringing the total staff to seven. When Rod accepted the Sheffield Chair there were plans for the Departments of Microbiology and Genetics to occupy a new Biology Building, but the plans were soon revised in favour of another Department. For the next seven years Microbiology was confined to the cramped conditions of the Elsden Department with no offices for staff and only four spaces to accommodate research students. There was a small office for the Professor and Secretary and just four small laboratories surrounding the chimney of the University incinerator, one of which was infested by rodents entering from the local park and escaping from the adjacent animal house.  John Guest remembers occupying two sides of an island bench in Rod's Lab and his desk being next to a large fermenter growing bacteria on a possibly explosive methane-air mixture with Peter White having a desk crushed into a closed doorway.
       Applying for research grants was a problem as there was no space for additional staff or equipment and in other respects these were frustrating times because Microbiology including Microbial Genetics and Molecular Genetics were expanding at an explosive rate, providing the materials and techniques for the genetic revolution and the emerging era of recombinant DNA.  The Microbiology Department seemed to be the only Department wanting to develop along these lines. Under Bartley the Biochemistry Department had grown in size but was little interested in Molecular Biology or in collaboration with other Departments. In his obituary of Krebs (8), written in 1980, Rod wrote “Krebs appealed to Sheffield University in 1951 to provide him with more space, arguing that the scant space given to the Biochemistry Department (⅙th of the space available to Chemistry and ⅓rd of that available each to Zoology and Botany) suggests that the general importance of Biochemistry as an academic subject has not been appreciated by the Sites and Building Committee". Rod goes on to write "Add 30 years on to 1951 and substitute Microbiology for Biochemistry, and you have a paragraph in current use by many Heads of British Microbiology Departments”. Eventually in 1972, dedicated teaching space for Microbiology and more space for research was acquired when another Department was relocated to the new building.
       Everyone in the Microbiology Department including Rod had a heavy teaching load. They continued to teach in the Biochemistry Degree course and in 1966 started a new Dual Honours Degree in Genetics and Microbiology in the outlying Genetics Department. Then, from 1976, they offered a Single Honours Degree in Microbiology in addition to Dual Degrees with Biochemistry, Genetics and ultimately with Biotechnology.  The courses included a wide range of topics reflecting the staff’s expertise in microbial structure, function, physiology and metabolism, growth kinetics and energetics, environmental microbiology, bacterial and phage molecular biology and molecular genetics, genetic engineering and biotechnology.
       Rod was an excellent Head of Department, always approachable and friendly to everyone.  By regularly attending weekly seminars he kept aware of the research of all the members of staff and their students. Decisions were made democratically by common consent at informal meetings of the academic staff over lunch on Fridays during term and in this way a harmonious and highly efficient Department was maintained.
With so many constraints of time, space and University attitudes it is truly remarkable how much was achieved by this small Department, not least the fact that two of the seven members of staff were elected Fellows of the Royal Society (J Rod Quayle FRS 1978 and John R Guest FRS 1986).

Bacterial growth on methane; the methanotrophs

After moving to Sheffield in 1963 Rod turned his attention to the methanotrophs – those bacterial methylotrophs that usually grow only on methane, but sometimes also on methanol. In their first paper on the serine cycle in methanol-utilisers, he had said that “It remains to be seen whether microbial growth on methane involves a metabolism broadly similar to that involved in growth on methanol” (9). This was particularly relevant to methanotrophs as it had previously been assumed that methane would be assimilated as carbon dioxide by the RuBP pathway of carbon dioxide fixation. 
        If he had chosen to study a different methanotroph he might have come to the obvious conclusion that all methylotrophs, including methanotrophs, would use the serine sycle for assimilating methane as well as methanol. But his first paper based on work at Sheffield (with Pat Johnson) showed that neither of the known pathways is operating (10). During growth on methane, ribulose bisphosphate carboxylasewas absent from Pseudomonas methanica (now called Methylomonas methanica), and the 14C-labelled compounds accumulating at early times during incubation with 14C-methane or 14C-methanol were neither phosphoglycerate nor serine but mainly glucose and fructose phosphates. Their demonstration of a novel formaldehyde-condensing enzyme in crude cell extracts indicated the presence of a pentose phosphate cycle for formaldehyde assimilation, analogous to the RuBP pathway for carbon dioxide assimilation. This was confirmed by a thorough examination of the labelling patterns and by characterising the novel enzyme responsible for condensing formaldehyde with ribulose monophosphate to give a sugar phosphate (with his student Michael Kemp). This sugar phosphate was later identified as a novel hexulose phosphate, D-arabino-3-hexulose phosphate (Kemp, 1974). Rod eventually completed the pathway, now known as the ribulose monophosphate pathway (RuMP pathway) by purifying and characterising a second novel enzyme, hexulose phosphate isomerase, and by demonstrating the presence of essential cleavage and rearrangement enzymes (for review see 4). In summary, 3 molecules of formaldehyde are condensed with RuBP to produce 3 molecules of hexulose phosphate which undergo a series of cleavage and rearrangement reactions to regenerate the acceptor ribulose monophosphate and to produce a three‑carbon phosphoglycerate for assimilation into cell material (Figure 3).


Figure 3. The ribulose monophosphate (RuMP) pathway of formaldehyde assimilation (KDGP aldolase / transaldolase variant). This cyclic pathway achieves the conversion of three molecules of formaldehyde to one molecule of phosphoglycerate which is the starting point for biosynthesis of all cell materials. The numbers refer to the enzymes catalysing the reactions. (8) transketolase; (9) pentose phosphate epimerase; (11) pentose phosphate isomerase; (12) transaldolase; (13) hexulose phosphate synthase; (14) hexulose phosphate isomerase; (16) glucsose phosphate isomerase; (17) glucose phosphate dehydrogenase; (18) phosphogluconate dehydrase; (19) 2‑keto, 3‑deoxy, 6‑phosphogluconate (KDPG) aldolase; (20) PEP synthetase or equivalent enzyme(s); (21) enolase; (22) phosphoglyceromutase. This Figure is reproduced with permission from Anthony (1982).  

       After this first description of the RuMP pathway in methanotrophs growing on methane, it was shown to be the main route for methanol assimilation in many bacteria including Methylophilus methylotrophus, the bacterium used by ICI for their SCP Pruteen project. This was significant because the RuMP pathway is energetically (and therefore commercially) more favourable than the serine cycle for bacterial growth on methanol. He also demonstrated that by having an extra enzyme (6-phosphogluconate dehydrogenase) in the pathway, some of these bacteria can oxidise formaldehyde to CO2 by a cyclic route instead of the usual linear pathway. He showed that there are four possible variants of the RuMP pathway, depending on which rearrangement reactions are operating (4).
Although Rod showed that the RuMP pathway operates in some other methanotrophic species he soon obtained evidence that at least one of them must use a different pathway. Short-term labelling experiments with Methanomonas methano‑oxidans identified serine and carboxylic acids as the early-labelled products as had been observed for methanol assimilation in Pseudomonas AM1, and key enzymes of the serine cycle were present during growth on methane whereas those of the RuMP pathway were absent. This provided the first indication that there are two metabolically distinct groups of methylotrophic bacteria. In 1970 another of the great milestones in C1 metabolism was reached when Roger Whittenbury, John Wilkinson and their colleagues transformed the field by their detailed study of elective culture of methanotrophs which resulted in the isolation and characterisation of over 100 new strains (Whittenbury et al, 1970). These formed two distinct groups according to the arrangement of their internal membranes, and Rod quickly confirmed that bacteria in one group used the RuMP pathway while the other used the serine cycle (11).

Methylotrophic yeasts

In the late 1970s Rod transferred his attention to some recently described methanol-utilising methylotrophic yeasts (4). In 1977 Hans van Dijken from the University of Gröningen was spending a year in Sheffield and Rod, together with other colleagues from Gröningen and Sheffield decided to examine carefully the literature suggesting that Candida uses the RuMP pathway during growth on methanol (12). They repeated the labelling experiments with 14C-methanol, using Hansenula polymorpha in place of Candida and obtained identical results for the two yeasts. However, cell‑free extracts of methanol-grown H. polymorpha and Candida boidinii lackedthe two key enzymes of the RuMP pathway, viz.  hexulose monophosphate synthase and hexulose phosphate isomerase, which were investigated using the authentic substrates and sensitive spectrophotometric assays that Rod had developed for the bacterial enzymes. Thus, although the 14C labelling experiments and the presence in cell-free extracts of an enzyme system capable of catalysing a pentose phosphate-dependent fixation of formaldehyde pointed towards a yeast equivalent of a bacterial RuMP pathway, the absence of essential key enzymes showed this is not the case. Roger Cox, a post-doctoral colleague, then drew their attention to the possibility that a pentose phosphate-dependent fixation of formaldehyde in crude extracts might be catalysed by transketolase, using xylulose 5-phosphate as the ketol donor, and formaldehyde as acceptor, a reaction that he had considered with Len Zatman during their investigation of the growth of Bacterium 2B2 on trimethylamine (Cox & Zatman, 1974). Based on such a condensation reaction, Rod and his colleagues proposed another novel assimilatory cycle, the dihydroxyacetone cycle for formaldehyde fixation in which dihydroxyacetone would be the primary product formed from formaldehyde (12). Elevated activities of a dihydroxyacetone kinase and fructose 1,6-bisphosphatase would be required during growth on methanol compared to with growth on multi-carbon substrates and this was confirmed in methylotrophic Hansenula and Candida species. It was then shown by Mary O’Connor (now Mary Lidstrom), a visitor from the USA, that mutants of H. polymorpha and C. boidinii lacking dihydroxyacetone kinase were unable to grow on methanol (13). In one respect at least Mary was typical of visitors to Rod’s lab, gaining techniques, experimental approaches and inspiration, and then over 35 years continuing to be an inspiration and leading light in the hugely expanded field of methylotroph genetics. 

Some further key questions answered

The timely identification of important key questions was a common feature of Rod’s approach, sometimes leading to extensive studies as in the assimilation pathways in bacteria and yeasts but sometimes involving a short successful foray into a related field. A good example of this is his work on methane oxidation with Dr John Higgins. Methane is rather inert chemically and it had been proposed that the first oxidative step in its metabolism would be catalysed by a monooxygenase, with energy being put into the reaction by the reductant NADH.  This was very difficult to confirm using conventional biochemical techniques but Rod saw an opportunity to solve the problem by using ‘heavy oxygen’ 18O, both as the gas and as a constituent of water. In 1970 with Higgins he showed unequivocally that the initial reaction must be catalysed by a monoxygenase, incorporating 18O into methanol from 18O2, but not from water containing 18O.
       Rod spent a sabbatical year as a visiting professor in Göttingen (1973-74), working with Professors Gerhard Gottschalk, Norbert Pfennig & Hans Schlegel at the famous Institut für Mikrobiologie in the Georg-August-Universitat.  Norbert Pfennig had shown that some members of the photosynthetic bacteria (the Rhodospirillaceae) are able to use methanol instead of carbon dioxide as their carbon source during anaerobic photosynthesis. During his visit Rod found a better way of isolating these bacteria and showed that it was likely (and now confirmed) that they assimilate methanol carbon after first oxidising it to carbon dioxide. Returning home he collaborated with a visitor from Germany, Hermann Sahm, and Roger Cox to confirm that the RuBP pathway is indeed used for growth on methanol after its initial oxidation to carbon dioxide. During this process methanol is oxidised by the NAD-independent quinoprotein methanol dehydrogenase which passes on its electrons to a special cytochrome c and thence, in aerobic conditions, to oxygen. However, Rod’s measurements of growth yields during anaerobic photosynthesis on methanol [14] showed that the dehydrogenase is also responsible for reduction of NAD to NADH, essential for assimilation of the carbon dioxide produced by methanol oxidation. This process must involve oxidation of the cytochrome c by a cytochrome bc1 complex, ubiquinone and NADH dehydrogenase. It is thus a rare example of ‘reversed electron transport’ from methanol in bacteria, the energy being provided by the light reactions of photosynthesis (see Anthony, 1982).

Industrial links and Conferences

In the late 1960s the discovery of the North Sea oil and gas fields led oil and chemical companies to realise that they had a new and cheap chemical and biological feedstock on their doorsteps. Accordingly in 1967 the Institute of Petroleum organised a Symposium in London on Hydrocarbon Microbiology. Both Rod and Douglas Ribbons had included methane microbiology in their talks and after the symposium Rod was approached by two chemists, P.P. King and D. Watchorn, from the Agricultural Division of Imperial Chemical Industries (ICI, at Billingham, Cleveland, UK) who felt that ICI might be interested in the possibility of very large scale microbial conversion of methane to bacterial protein for use as an animal foodstuff.  They invited Rod to Billingham where in discussion he persuaded them that methanol would be a far more suitable substrate than methane.  P.P. King recalls this “As a Eureka situation" adding that "if there is one thing we can do it is to make methanol out of natural gas very efficiently”. Out of these beginnings the ICI Pruteen project was born.  Within the astonishingly short time of 13 years from the first discussions, the world's largest fermenter was constructed for the fast-growing Methylophilus methylotrophus, and full production was achieved on Teesside in 1980 (Figure 4). The Pruteen output from the 50m high (1.5 million litre) airlift fermenter was 50,000 metric tons per annum. Sadly this pioneering ICI project costing well in excess of £100 million had to be abandoned due to falling prices of competing products such as soybean protein.  Although nutritionally excellent, Pruteen was just too expensive to produce. However, thanks to his initial relationship with ICI, this company was for many years a very welcome sponsor of research by many academic microbiologists in the UK engaged in fundamental work on methylotrophic bacteria. This quest for inexpensive single-cell protein (SCP) was a world-wide phenomenon and once the industrial giants like ICI, BP and Shell had become interested in methane and methanol microbiology, large research funds began to flow, and the study of methylotrophs rapidly gathered pace in Europe and the USA, with research groups forming in Sittingbourne, Canterbury, London, Hull, Grangemouth, Gottingen, Madison, Dallas, Puschino, Kiev, Tokyo, Kyoto and Haren, in addition to those already active in Sheffield, Edinburgh and Reading.
        By 1973, there was sufficient interest for Rod, together with John Wilkinson and Roger Whittenbury, to raise financial support from BP, ICI and Shell to organise in Edinburgh what must be considered as the first authentic C1 Symposium. There were 50 participants and it lasted 2½ days. The first ‘official’ International Symposium on The Microbial Growth on C1 Compounds was held a year later in Tokyo and this was followed every three years by wonderful symposia, in Puschino, Sheffield, Minneapolis, Haren, Gottingen and Warwick. The eighth of these was held in San Diego in 1995 and since then they have continued every two years under the auspices of the Gordon Research Conferences, the most recent being in 2014. It was at these meetings that Rod’s influence was most personally experienced. His authority coupled with his modesty and helpfulness made his contributions eagerly anticipated. His lecturing style was rather formal, often speaking from a typescript, whilst his informal contributions often took the form of elegant hilarious anecdotes often aimed at the pompous or self-important. He will be remembered by those who knew him at these conferences as the voice of reason, a serious intellect, generous in his advice and help, bringing a compassionate almost genteel understanding of anyone’s problems, personal or scientific. His valedictory lecture at the 1995 symposium on Microbial Growth on C1 Compounds in San Diego was typical of the man in which he highlighted all the achievements since the 1st Symposium 22 years earlier and played scant attention to his own discoveries even though this had influenced nearly every facet of C1 metabolism for over 30 years.

Figure 4. The ICI ‘Pruteen’ plant at Billingham on Teeside, UK. The specially developed airlift fermenter is the middle tower; the others are for the production of growth medium and for cooling.

Other aspects of his time in Sheffield

In 1974 it became Rod’s turn to serve as Dean of the Faculty of Science (1974-1976) during which he was able to award the first Krebs Prize for Biochemistry on Degree Day.  He had established this prize by generously using the royalties accruing from a book of essays by former colleagues of Krebs and dedicated to Krebs on his 70th birthday. He also established the Boehringer Prize for the best Microbiology graduate.  In this case the original fund came from a substantial sum of money received by the Microbiology Department from the Boehringer Chemical Company as a discount based on their huge consumption of coenzymes and other fine chemicals. The money was intended to be used to buy further Boehringer chemicals, but Rod asked for it to be used to create a Prize in Boehringer's name and the Company agreed.
       Rod’s extraordinarily fruitful exploration of the biochemistry of the highly diverse range of microbes growing on C1 compounds during his time in Oxford and this period in Sheffield led in 1978 to the award of the CIBA Medal and Prize of the Biochemical Society (4), and his election as a Fellow of The Royal Society. If he had ‘merely’ elucidated a single novel pathway for microbial carbon assimilation, that pathway would probably have become the ‘Quayle Cycle’. This could not happen because he had described three novel pathways: the serine cycle, the ribulose monophosphate pathway (and its four variants) and the dihydroxyacetone pathway.
       His time in Sheffield was shared with his wife, Yvonne, and their 2 children, Susan and Rupert. When Rod organised a highly successful C1 Symposium (1980) in which about twenty five nations were represented, Yvonne organised the social activities for accompanying wives. She often joined him on his visits to the C1 Symposia, lightening and leavening the social aspects of the science community. She became an accomplished painter and an active member of the University Wives’ Club and remembers their stay in Sheffield period as a great time of entertaining. With three friends Yvonne produced an excellent cookery book - 'Cooking by Degrees' containing recipes born of Rod's enthusiasm for bread-making: 'Quayle Bread' and 'Rod's Whole Meal Bread'.
       Rod was always very much a family man - enthusiastically wrapped up in his two children and family activities.  His leisure activities were pretty simple, a major relaxation being walking in the Peak District, right on his doorstep.

BATH (1983 – 1992)

Twenty years after moving from Oxford to Sheffield Rod felt he had achieved as much as he could in the areas in which he was interested and after considering a number of other options he moved to the University of Bath as Vice-Chancellor where it is likely that his various experiences at Sheffield had an important influence in his attitudes and approaches to University organisation. 
       This was a time of significant change in the restructuring of higher education management and performance in the United Kingdom, so Rod had to adapt to the role of businessman and chief executive. He had those qualities of making things happen, making people understand and follow his objectives, whilst at the same time getting on and steering the ship. Sometime after the Jarratt Report of 1985 he was charged by the University Council to implement, within three years, changes in the University organisation. This he achieved so successfully that within one year the University of Bath was transformed from a highly centralised managed university to one with devolved management and executive authority devolved to heads of departments. During the nine years under his leadership Bath University rose to be an outstanding research university with 11 of the 14 Schools of Study ranked within the top grade for UK universities in the Research Assessment Exercises.
       During his time at the University of Bath, he vigorously championed this relatively young university as a centre for both pure and applied research, in the conviction that there is only one Science -"Science applied to useful ends" and "Science waiting to be so applied".  The remarkable growth in size, scope and influence of the University of Bath owes much to Rod's vision, leadership and enthusiasm for science. In spite of his involvement in all aspects of the life of the University Rod maintained his enthusiastic interest in the research of all those within his own field of interest - the microbial metabolism of C1 compounds. Even when unable to attend the international conferences, he was always eager to hear reports of recent advances and was always available to accept visitors who knew that time spent with him would revitalise their own enthusiasm. Here a personal anecdote: a visitor to Southampton from the then repressive East Germany (Wolfgang Babel) asked me if it would be possible to meet his hero and inspiration. After one telephone call, a visit was set up for the very next day.  It was spent in very animated discussion and debate, almost leaving my visitor in tears and saying “I hope you in the UK appreciate what a sensible system you have that gives power and influence to such a great, wise and generous man as Professor Quayle”. A measure of the personal respect, admiration and affection in which Rod was held by all those working on methylotrophs can be found in the preface to The Biochemistry of Methylotrophs (Anthony, 1982) dedicated to J. Rod Quayle, ‘The Godfather of Methylotrophy’.
       Rod’s relationship with students was quite special. He had a way of talking and always listening to students’ needs. Typically he assumed that it was his place to offer lifts in his car to students who waited at the bottom of the long hill leading from the town to the campus. Although trying to avoid it he sometimes had to admit that he was the Vice-Chancellor but their shocked response was so great he eventually he gave up the practice as being unkind.
His continuing work with the Royal Society brought about one of the coups of the time by transferring the National Cataloguing Unit for Archives of Contemporary Scientists to its location in the University of Bath.  He was also influential and involved in bringing together the combined strengths of the Universities of Bristol, the West of England and Bath, in the planning and initial strategy for a major science park.  Working on the national scene, he was elected Chair of the UK National Committee for Microbiology (1985 -1990), advising the UK government on all microbial matters, and he served as President of the Society for General Microbiology from 1990 to 1993. He continued to work in various capacities in the Royal Society until a few months before he died.
       While working on behalf of the University of Bath Rod also played a part in the local community and in national and international affairs, both academic and social. These included the reformation of the Bath Royal Literary and Scientific Institution whose aim was the “Promotion and Advancement of Science, Literature and Art”. He was a member of the Council of the Bath Institute of Medical Engineering which “Uses the multidisciplinary approach of medicine, engineering and science to identify needs of disabled people and hospital patients not being met elsewhere and to provide solutions”. He was a member of the Board of the Bristol Exploratory, a hands-on Museum of Science founded by Richard Gregory. He also made a significant contribution to one of Yvonne’s and his great loves in the world of music and the arts, by inspiring the management of the world-famous Bath Festival, which provided a happy, relaxed reward for them when dutifully meeting the performers and attending great concerts.

RETIREMENT (1992 – 2006) AND FAMILY

The first thing that bothered Rod in retirement was that he would no longer have a secretary, so he had to learn how to use a computer.  At first he was very busy as President of the Society for General Microbiology.  He also wrote Biographical Memoirs for Fellows of the Royal Society for W. Charles Evans (15) who had been a Professor of Biochemistry at Bangor, and Leonard Rotherham (16) an engineer who had been a previous Vice-Chancellor of the University of Bath.
       During retirement, in the small Somerset village of Compton Dando, Rod continued with many of those activities he had enjoyed as Vice-Chancellor at Bath, his hobbies including walking, swimming and going to concerts and the theatre in Bristol and Bath. He also had more time for cooking, baking bread and gardening.
      Yvonne and Rod’s daughter, Susan, went to Durham University to read Law, qualified as a solicitor and now works in Bath as the in-house lawyer for a publishing company with various overseas offices. Their son Rupert did 4 years’ training at Grimsby Nautical College and has since enjoyed a career as a Captain in the Merchant Navy. Susan has three children and Rupert has two. Rod always played an active role in looking after the five grandchildren: taking them for days out, to swimming lessons, sometimes picking the local ones up from school, and watching school plays. All the grandchildren are very fond of each other - and they too were very fond of Rod. He loved small children and they loved him. As well as being a born raconteur, Rod also had an infectious sense of humour which is still remembered by everyone who knew him.

HONOURS

As mentioned in context above, in 1978 Rod was awarded the CIBA Medal and Prize of the Biochemical Society in honour of his outstanding contribution to Biochemistry (4). In the same year he was elected as a Fellow of The Royal Society in which he served as Chair of one of the Sectional Committees and also as a Member of Council (1982-1984). From 1990 to 1993 he was President of the Society for General Microbiology. During a 3 month visit as visiting professor of Microbiology in the University of Washington at Seattle he received the Walker-Ames medal. He was awarded honorary doctorates from the Universities of Göttingen (1989), Bath (1992) and Sheffield (1992), and had an Honorary Fellowship conferred upon him by Bangor University in 1996.

ACKNOWLEDGEMENTS

I should like especially to thank John Guest (FRS 1986) for his hard work and enthusiastic involvement in every aspect of the production of this memoir. Professor Sir Hans Kornberg (FRS 1965) provided important information on Rod’s work in California and Oxford, and Richard Mawditt was especially helpful about Rod’s time as Vice-Chancellor of the University of Bath. I am personally grateful to his widow, Yvonne, and their daughter, Susan, for invaluable information and enjoyable reminiscing. Finally I wish to record my gratitude to the Royal Society for giving me the opportunity to write this memoir of a great microbiologist.

REFERENCES TO OTHER AUTHORS

Anthony, C. 1982 The Biochemistry of Methylotrophs.  Academic Press: London, 430 pages.

Anthony, C. 2011 How half a century of research was required to understand bacterial growth on C1 and C2 compounds; the story of the serine cycle and the ethylmalonyl-CoA pathway. Science Progress, 94, 109–137.

Cox, R. B. & Zatman, L. J. 1974 Hexose phosphate synthase in trimethylamine-grown bacterium 2B2, a facultative methylotroph. Biochem. J. 141, 605–608.

Kemp, M. 1974 Hexose Phosphate Synthase from Methylococcus capsulatus makes D-arabino-3-hexulose phosphate. Biochem. J. 139, 129-134.

Kornberg, H. L. & Krebs, H. A. 1957 Synthesis of cell constituents from C2-units by a modified Tricarboxylic acid cycle. Nature 179, 988–991.

Whittenbury, R., Phillips, K. C. & Wilkinson, J. F. (1970a). Enrichment, isolation and some properties of methane-utilizing bacteria. J. Gen. Microbiol. 61, 205-218.

  TOP                                              BIBLIOGRAPHY

The following publications are those referred to directly in the text. A full bibliography is available as a pdf

(1)           1961      Metabolism of C1 compounds in heterotrophic and autotrophic micro-organisms.     Ann. Rev .Microbiol. 15, 119-152.

(2)           1972      The Metabolism of One-Carbon Compounds by Micro-Organisms. Adv. Microbial Physiol. 7, 119-203.

(3)           1978      (With T. Ferenci) Evolutionary aspects of autotrophy. Microbiological Reviews 42, 251-272.

(4)           1980      Microbial assimilation of C1 compounds. Thirteenth CIBA Medal Lecture. Transactions of the Biochemical society 8, 1-10.   

(5)           1954      (With R.C. Fuller, A.A. Benson & M. Calvin) Enzymatic carboxylation of ribulose diphosphate. J. Amer. Chem. Soc. 76, 3610.

(6)           1958      (With H.L. Kornberg) The metabolism of C2 compounds in micro-organisms. 2. The effect of carbon dioxide on the incorporation of 14C acetate by acetate-grown Pseudomonas Kb1.  Biochem. J. 68, 542–549.

(7)           1962      (With P.J. Large & D. Peel) Microbial growth on C1 compounds. 3. Distribution of radioactivity in metabolites of methanol-grown Pseudomonas AM1 after incubation with [14C] methanol  and [ 14C] bicarbonate.  Biochem. J. 82, 483-488.

(8)           1982      Obituary. Sir Hans Krebs, 1900-1981. J. Gen. Microbiol. 128, 2215-2220.

(9)           1961      (With P.J. Large & D. Peel) Microbial growth on C1 compounds. 2. Synthesis of cell constituents by methanol- and formate-grown Pseudomonas AM1, and methanol-grown Hyphomicrobium vulgare. Biochem. J. 81, 470-480.

(10)        1965      (With P.A. Johnson) Synthesis of cell constituents by methane- and methanol‑grown Pseudomonas methanica. Biochem. J. 95, 859-867.

(11)        1970      (with A.J. Lawrence) Alternative carbon assimilation pathways in methane-utilising bacteria. Journal of General Microbiology, 63, 371-374.

(12)        1978      (With J.P. van Dijken, W. Harder & A.J. Beardsmore. Dihydroxyacetone: an intermediate in the assimilation of methanol by yeasts? FEMS Microbiology Letters 4, 97-102.

(13)        1978       (with M.L. O’Connor) Mutants of Hansenula polymorpha and Candida boidinii impaired in their ability to grow on methanol. J. Gen. Microbiol. 113, 203-208.

(14)        1975      (With N. Pfennig) Utilisation of methanol by Rhodospirillaceae. Arch. Microbiol. 102, 193-198.

(15)         1994      William Charles Evans Biographical Memoirs of Fellows of the Royal Society 40, 87‑103.

(16)         2003      (With G.W. Greenwood) Leonard Rotherham. Biographical Memoirs of the Royal Society 49, 433-446.

Powerpoint presentation of a lecture 'In Memoriam Rod Quayle' given at a Gordon Conference in Oxford in

Obituary published in the Biochemist