SCIENCE   General
                  My research


INDIA      General
               My pictures  

FAVOURITES shopping, friends,
DIY, entertainment,
Southampton etc







   All engines


NOTE 'My files' open with:
ppt Powerpoint
doc Word
xls Excel
pdf Acrobat Reader      [download free]
jpg Opens direct
html Opens direct;
For BIGGER text
Click Ctl plus +
Normal text click Ctl plus 0

VIDEOS can be speeded up by downloading an accelerator




Return to: Research and teaching page
A History of Methanotrophy

Methylotrophic bacteria are bacteria that are special in that they can grow on compounds with only one carbon atom, such as methane, methanol, methylamine. All their cell carbon must be synthesised from these compounds and all their energy must be obtained by their oxidation.

Methanotrophs are the most specialised of the methylotrophic bacteria because they are able to grow on methane (and sometimes methanol). The great majority of these bacteria are obligate methanotrophs, unable to grow on multicarbon compounds.

Because methane is relatively inexpensive it is potentially important as a primary source for the production of chemicals by methanotrophs on an industrial scale.

Methane is an important greenhouse gas (18 times more 'effective' than carbon dioxide). The methanotrophs play an important role in the carbon cycle in nature, removing it methane by growing on it.

Because of a growing appreciation of its importance a special website has been established where information and opportunity for discussion can be provided.
This website is called Methanotroph Commons. I have prepared a History of Methanotrophy for this website and have inlcluded the same material here on my website.

For extensive sets of slides illustrating most of the topics covered here, click: Powerpoint presentations. Some topics are also covered in detail in summaries of my research
The contribution of J. Rod Quayle to the subject is also covered in a Biographical Memoir. At the end of each section there is a direct link to a set of relevant illustrations, including pictures of some of the scientists involved.

Thanks to Prof. Sai Gopal and Sri Venkateshwara University and their Guesthouse in Tirupati where this History was written [pictures of this wonderful place]

The Methanotrophs; a History         

By Chris Anthony, University of Southampton, Southampton, SO17 1BJ,  UK.
c.anthony@soton.ac.uk;   chris-anthony.co.uk

Download this History as a Word file       Return to My research and teaching topics

This history is in six sections:

Introduction;             Carbon assimilation;       Methane oxidation;   
Methanol oxidation
;    Electron transport;         Genetics.

Of course, most reviews of any aspect of the subject will include some historical context. What I have tried to do is to be as thorough as possible when covering the beginnings. If there are any omissions, errors or insults please contact me to correct them. Much of the biochemistry of methanotrophs was first elucidated in methylotrophs growing on methanol, but unable to use methane. I will concentrate on methanotrophs when possible but use work with other methylotrophs where appropriate. Much of the history of each subject is covered in detail in  my book The Biochemistry of Methylotrophs (1982) which covers the subject up to 1982 in 400 pages with more than 1000 references. This is not available in print but it can be downloaded as pdf from my website: http://www.chris-anthony.co.uk/methylotrophs.html.   I own the copyright so downloading is free; help yourself.


Methanotrophs, growing on methane as their sole source of carbon and energy, are a specific category of the more general category of Methylotrophs which grow on carbon compounds containing one or more carbon atoms but with no carbon-carbon bonds (eg methane, methanol, methylamine, trimethylamine). Methanotrophs are almost always obligate, growing only on methane or (sometimes) methanol which is the first intermediate in methane utilisation. All methylotrophs share two common challenges: they must obtain energy in the form of ATP and NAD(P)H by oxidation of their unusual substrates, and they must have special pathways for carbon assimilation, as all their carbon-carbon bonds must be made de novo.

The ‘historical’ start of published work on methanotrophy is usually taken as the limited description in 1906 of an organism isolated on methane from aquatic plant material by Sohngen (1906, 1910) and named by him Bacillus methanicus (later amended by Orla-Jensen (1909) to Methanomonas methanica). It was only able to grow on methane or methanol. A firmer foundation was provided 50 years later by Dworkin and Foster (1956) with the full description and classification of “a pink methane-utilizing bacterium in all respects identical with Methanomonas methanica Sohngen” which they showed to be widely distributed in nature. They considered that “the genus Pseudomonas accommodates the methane-utilizing bacterium here under consideration perfectly satisfactorily” as it is “an aerobic, gram negative, polarly monoflagellated rod”. They proposed, therefore, “to rename Sohngen's organism Pseudomonas methanica (Sohngen) nov. comb”. “The distinctive physiological character of this species is its extremely limited substrate range: it cannot use any of a large number of conventional carbon sources and methane is the only saturated gaseous hydrocarbon which it can oxidize”.

They rejected the name Methanomonas because the prefix Methano- was widely used for methanogenic bacteria. They then argued that “From the viewpoint of modern microbiology and systematics, generic designation on the basis of the utilization of particular compounds as a sole source of organic nourishment would hardly be tactful. If adopted in the case of all organic compounds arbitrarily regarded as unusual substrates, such a practice would produce taxonomic and nomenclatural chaos”. They considered that “recognition of physiological genera based on methane utilization would confer no advantage on systematics or diagnostics, and would imply a phylogenetic distinction more profound than is warranted. Furthermore, it might be a precedent for a whole series of trivial genera in connection with other individual compounds”. With hindsight of course, in the case of methane and methanotrophs this sensible caution and tactfulness has proved to be unnecessary (note: Pseudomonas methanica is now known as Methylomonas methanica). In the next fourteen years a few more methanotrophs were described, all being strictly aerobic, and able to grow on methane and methanol but not on multicarbon compounds; these included Methanomonas methanooxidans (Brown et al., 1964) and Methylococcus capsulatus (Foster and Davis, 1966).

A major landmark in the development of our knowledge of methanotrophs was the description in 1970 of more than 100 new isolates by Whittenbury, Wilkinson and their colleagues at Edinburgh (Whittenbury et al., 1970a,b). All were obligate methylotrophs able to grow only on methane (and sometimes methanol), but they were very diverse with respect to biochemistry and structural characteristics. This, together with the observation that no previously-described species of bacteria had been shown to grow on methane, made it obviously acceptable to discard Dworkin and Foster’s ‘tactful’ approach and to give the five ‘groups’ of isolates names that indicate their physiological and nutritional speciality, and these groups have subsequently been accepted as the genera Methylocystis, Methylococcus, Methylosinus, Methylomonas and Methylobacter.   Some strains were considered to be identical with previously described methane-utilizing species, such as Pseudomonas methanica (Dworkin & Foster, 1956) and Methylococcus capsulatus (Foster & Davis, 1966). But none was identified as Methanomonas methanooxidans (Brown et al., 1964).
TOP of Introduction       References for Introduction        TOP of Pages

Illustrations of the Methylotrophs and the scientists involved

Carbon assimilation pathways                        TOP of Pages  

Ten years before Whittenbury’s great contribution to the study of methanotrophs, an extensive programme of study of methylotrophs growing on methanol was initiated by J. Rod Quayle in the University of Oxford (to be continued in the University of Sheffield, UK). He soon became established as an inspirational leader in the study of methylotrophs. He had worked in Calvin’s lab on the path of carbon in photosynthesis, and had published the first demonstration of the “Enzymatic carboxylation of ribulose diphosphate” by Chlorella extracts (by what we now know as ribulose bisphosphate carboxylase/oxygenase; RuBisCo) (Quayle et al., 1954). In 1961, with David Peel, he published a description of Pseudomonas AM1 (Peel & Quayle, 1961) which has become the ‘work-horse’ of methylotrophy, It  is a pink facultative methylotroph that does not grow on methane but grows on methanol, methylamine, formate and also on multicarbon compounds. It is now called Methylobacterium extorquens AM1; other earlier names have included  Pseudomonas AM1, Pseudomonas sp. M27, Vibrio extorquens and Pseudomonas extorquens.

It had long been thought that reduced C1 compounds might first be oxidised to carbon dioxide and then assimilated by the Ribulose bisphosphate cycle or ‘Calvin cycle’as in plants and autotrophic bacteria , and Quayle had confirmed that this could be a possibility by showing that formate is assimilated by this cycle in Pseudomonas oxalaticus (Quayle & Keech, 1959a,b). However, that serine rather than phosphglycerate might be a possiblefirst intermediate during methanol assimilations in a pink facultative methylotroph Pseudomonas PRL-W4 (similarto Pseudomonas AM1) had been suggested by Kaneda & Roxburgh (1959a,b). Subsequently, using the experimental approaches that had led to the elucidation of the ‘Calvin cycle’, Peter Large, David Peel and Rod Quayle, in a series of model papers, showed that methanol is assimilated in Pseudomonas AM1 at the levels of formaldehyde and carbon dioxide by a novel biosynthetic cycle, the Serine cycle (Large et al., 1961; 1962a,b; Large & Quayle, 1963). At this time a key step in the pathway was missing which, ten years later, was identified by Salem & Quayle (1973) as a cleavage reaction catalysed by malyl-coenzyme A lyase which produced acetyl-Coenzyme A. A final step was to elucidate the  route for oxidation of this product to glyoxylate and the elucidation of this part of the pathway required almost another 40 years (see Anthony, 2011).

In their first paper on the Serine cycle in methanol-utilisers, Quayle and colleagues said that “It remains to be seen whether microbial growth on methane involves a metabolism broadly similar to that involved in growth on methanol” (that is, not involving the RuBP pathway of carbon dioxide fixation) (Large et al., 1961). That this was a possibility was indicated by the observation by Leadbetter & Foster (1958) that during growth of Pseudomonas methanica on methane a considerable proportion of the carbon incorporated by the cell does not pass through a stage readily exchangeable with carbon dioxide making it “unlikely that an autotrophic type of metabolism is involved”.

That a novel pathway was indeed operating during growth on methane in Pseudomonas methanica was confirmed by showing that ribulose bisphosphate carboxylase was absent, and that the 14C-labelled compounds accumulating at early times during incubation with 14CH4 or 14CH3OH were mainly glucose and fructose phosphates, and not the first product of the ribulose bisphosphate pathway of CO2 fixation (phosphoglycerate) (Johnson & Quayle, 1965). A formaldehyde-condensing enzyme was then demonstrated in crude extracts of Methylomonas methanica (previously called Pseudomonas methanica) by Kemp and Quayle (1965,1966), suggesting a novel pentose phosphate cycle for formaldehyde assimilation, in which the product of condensation with formaldeyde was tentatively identified as allulose phosphate (Kemp & Quayle, 1966). Thorough examination of the labelling patterns confirmed the proposed pathway (Kemp & Quayle, 1967). The pentose phosphate was subsequently shown to be ribulose monophosphate and the product of the condensation reaction is the novel sugar hexulose 6-phosphate) (Kemp, 1974). This pathway is now known as the ribulose monophosphate pathway (RuMP pathway).

That this, or a similar, pathway also operates in Methylococcus capsulatus, was shown by demonstrating that 14CH3OH was rapidly incorporated into hexose phosphates, and that an enzyme system is present which is able to condense formaldehyde with pentose phosphate to give a mixture of hexose phosphates (Lawrence et al., 1970).

The Ribulose monophosphate cycle of formaldehyde fixation (RuMP pathway) in methanotrophs was eventually completed by the purification and characterisation of the hexulose phosphate synthase and isomerase from Methylococcus capsulatus (Ferenci et al., 1974); and by demonstrating the presence of essential cleavage and rearrangement enzymes in Methylomonas methanica and Methylococcus capsulatus (Strom et al., 1974).

That all methanotrophs do not assimilate methane by the RuMP pathway was shown using the same short-term labelling techniques as were used with Methylococcus capsulatus but using Methanomonas methano-oxidans (a type II methanotroph – see below) (Lawrence et al., 1970). In this case, early label was found in serine and carboxylic acids (rather than in sugar phosphates) as had been found during methanol assimilation in Pseudomonas AM1. Furthermore, a key enzyme of the Serine cycle (hydroxypyruvate reductase) was present at high specific activity while the key formaldehyde-fixing enzyme of the RuMP pathway (hexulose phosphate synthase) was absent.

The presence or absence of these two key enzymes was subsequently used for allocation of putative pathways for methane assimilation by Whittenbury’s new isolates. Lawrence & Quayle (1970) showed that division of the organisms into two classes based on their assimilation

TOP of Carbon Assimilation pathways    References for assimilation
Illustrations of the pathways     Top of Page

Methane oxidation                                           TOP of Pages 

The most obvious suggestion for a system for oxidation of methane is that a monooxygenase is involved, catalysing this reaction:

CH4 + NADH + H+ + O2 CH3OH + NAD+ + H2O

That such a mixed function monooxygenase is involved was first implied by the work of Leadbetter & Foster (1959) on Pseudomonas methanica oxidising ethane and other alkanes (they did not investigate methane itself). This was later confirmed by Higgins and Quayle (1970) who showed that whole cells of Methylomonas methanica and Methanomonas methanooxidans incubated with methane incorporate 18O into methanol from 18O2 but not from water containing 18O. Further understanding of the nature of the methane monooxygenase (MMO) has depended on isolation of cell-free systems able to catalyse methane oxidation. This first depended on measuring NADH oxidation and O2 consumption in the presence of methane by particles prepared from Methylococcus capsulatus (Ribbons and Michalover, 1970) but as these preparations also oxidise NADH, in the absence of methane (Ribbons, 1975), interpretation presented considerable difficulties.

These difficulties were eventually overcome by John Colby, Howard Dalton and colleagues at Warwick who developed alternative assay methods that depended on the wide substrate specificity of the MMO; they measured oxidation of bromomethane or ethene or propene, their products being analysed by gas-liquid chromatography (Colby et al., 1975, 1977). This led them to provide the first definitive description of a soluble NADH-requiring methane monooxygenase, consisting of three proteins, present in Methylococcus capsulatus and in Methylosinus trichosporium OB3b (Colby & Dalton, 1978,1979) . This work has led to much of our modern understanding of methane oxidation in methanotrophs and is summarised in Howard Dalton’s Leeuwenhoek Lecture: The Natural and Unnatural History of Methane-Oxidizing Bacteria (Dalton, 1975), and in a  published tribute to Howard after his death (Anthony, 2008).  

That acquiring an understanding of methane oxidation in methanotrophs was not straightforward, is illustrated by the work of John Higgins and his group at Canterbury (Kent) who described a completely different particulate system from Methylosinus trichosporium which was said to use ascorbate or reduced cytochrome c, but not NADH, in solubilised, purified preparations (Tonge et al., 1977). However, Stirling & Dalton (1979) in a comparison of substrate and electron donor specificities in Methylococcus capsulatus, Methylomonas methanica and Methylosinus trichosporium concluded that NAD(P)H is the electron donor. The debate was further confused by difficulties in repeating experiments, some of the conflict being related to the subsequent observation that whether or not soluble or particulate MMO is produced is determined by the growth conditions. It is now known that copper stress underlies the intracellular location of MMO (Stanley et al., 1983). The soluble, NADH-requiring sMMO is produced when copper/biomass ratio is low, and the (completely different) membrane pMMO is produced when this ratio is high. It is now thought that the electron donor to the membrane enzyme (pMMO) is probably not NADH but membrane ubiquinol which may be produced by reverse electron transport from methanol by way of methanol dehydrogenase, its electron acceptor cytochrome cL and the cytochrome bc1 complex, driven by the protonmotive force provided by the oxidation of cytochrome cL. The details of the early confusing story of the development of understanding of methane oxidation are more fully covered by Anthony (1982).

Top of methane oxidation      References for methane oxidation
Illustrations for oxidation of methane, methanol and formaldehyde
TOP of Pages

Methanol oxidation                                         TOP of Pages 

The first step in the oxidation of methane is its oxidation to methanol which consumes energy in the form of NADH or ubiquinol. This must be provided by the subsequent oxidation of formaldehyde or formate, produced by the oxidation of methanol. Methanol is oxidised to formaldehyde by an unusual methanol dehydrogenase. During methane and methanol all carbon passes through this enzyme to produce formaldehyde which is either assimilated or oxidised.

I hope I will be forgiven for making the description of this part of the history of methanotrophy relatively personal. In 1960 I started working for my PhD in the Microbiology Department of the University of Reading, UK with Len J. Zatman as supervisor. He had previously worked on NAD-linked alcohol dehydrogenases and had been impressed that none of these is able to oxidise methanol. Hearing that some bacteria can grow on methanol he concluded that they must have an unusual enzyme to oxidise their growth substrate, and to find this enzyme was my PhD project. In my youthful ignorance I thought this might not be sufficiently interesting to occupy me for 3 years. But our first paper on the unusual methanol dehydrogenase was published in 1964 (Anthony & Zatman, 1964b) and my last paper, published on the atomic resolution of the same enzyme, was more than 40 years later (Williams et al., 2005). This history is dedicated to the memory of my hero and friend Len Zatman.

I started my project by isolating a pink facultative methanol-utilising (Pseudomonas sp. M27; Anthony & Zatman, 1964a). A year after we started, a description of a very similar organism was described by Peel & Quayle (1961) (Pseudomonas AM1).

The first reported investigation of methanol oxidation, in a similar organism, was by Kaneda & Roxburgh (1959a) who described methanol-dependent reduction of NAD by extracts of Pseudomonas PRL-W4, but this has never again been demonstrated. Harrington & Kallio (1960) subsequently suggested that methanol is oxidised in the methanotroph Pseudomonas methanica (Iowa strain) by the peroxidative action of catalase with methanol as substrate. Evidence against this was obtained from our inhibitor studies with methanol-grown Pseudomonas M 27 (Anthony & Zatman, 1964a). Complete inhibition of catalase activity had no effect on methanol oxidation; and conversely, methanol oxidation was completely inhibited by 5 mM-EDTA and the phenylhydrazines (0.1 mM) which had no effect on catalase activity. Harrington & Kallio (1960) suggested that the inhibition of methanol oxidation in whole cells by EDTA was due to inhibition of the iron-requiring catalase, this inhibition being relieved by addition of ferrous sulphate. But we showed this was an artefact caused by the aerobic inorganic chemical oxidation of Fe2+ to Fe3+ ion which occurs more rapidly in the presence of EDTA (Anthony & Zatman, 1964a). It was much later shown that EDTA inhibits methanol oxidation in whole cells by competing with the electron acceptor cytochrome cL for the same binding site on the methanol dehydrogenase (Chan & Anthony, 1992).  

Cell extracts of Pseudomonas M27 had, as expected, no NAD(P)H-linked methanol dehydrogenase activity. Eventually we demonstrated methanol-dependent reduction of the redox dye 2,6-dichlorophenolindophenol in the presence of an essential mediator phenazine methosulphate, using whole cells incubated anaerobically in Thunberg tubes. Initially this activity was not seen in spectrophotometric measurements with extracts because lab buffers contain tiny amounts of alcohols and so the rates were the same with or without added substrate. Furthermore, activity was never seen when smaller amounts of enzyme were used because we showed later that  the dye-linked methanol dehydrogenase requires ammonia (base) as activator, provided by addition of ammonium salts (Anthony & Zatman, 1964b).

The dye-linked methanol dehydrogenase (MDH) was soluble; it had a wide substrate specificity, was inhibited by none of the usual inhibitors, was very stable with a high isoelectric point and a molecular weight of about 120 kDa, being made up of 2 identical subunits of about 60 kDa (Anthony & Zatman, 1965, 1967a). Neither we ourselves, nor the dozens of later studies of MDH saw the other two tiny (8.4 kDa) subunits at that time; we found these more than 20 years later (Nunn et al., 1989). MDH was not very active which meant that a lot of enzyme had to be prepared for any study; but it also meant that, fortunately, the bacteria made a lot of it (about 10% of the soluble protein). Johnson & Quayle (1964) showed that the same enzyme is present in the very closely related Pseudomonas AM1 (Methylobacterium extorquens AM1).

Methanotrophs have the same (or similar) MDH as in Methylobacterium extorquens, as first demonstrated in Methylomonas methanica  by Johnson & Quayle (1964), The MDH was later purified and characterised from Methylococcus capsulatus and shown to be almost identical to that in Methylobacterium extorquens (Patel et al., 1972, 1973; Wadzinsky & Ribbons, 1975) and the MDHs from M. extorquens and methanotrophs (Methylococcus, Methylobacter and Methylosinus) were shown to be immunologically similar (Patel et al., 1973). The most different of the MDHs was that from Methylosinus sporium which was purified and crystallized by Patel & Felix (1976) who showed that it was less stable at low pH values and appeared to be monomeric (molecular weight 60kDa).

In 1967 we showed that MDH has a novel (unidentified) prosthetic group (Anthony & Zatman 1967b) which we purified (as a tiny amount of brick-red powder) and characterised by its fluorescence characteristics. We found out later that the same prosthetic group had been described in glucose dehydrogenase in Acinetobacter calcoaceticus by Hauge (1964). It was thirteen years after we first described the novel prosthetic group of methanol dehydrogenase that its structure was determined, by X-ray crystallography in Olga Kennard’s lab at Cambridge, to be pyrroloquinoline quinone (PQQ) (Salisbury et al., 1979). Methanol dehydrogenase was the first extensively studied example of what have become known as quinoproteins, a major new category of bacterial enzymes responsible for the first step in the oxidation of their growth

substrates. (For reviews of this enzyme and the many related quinoproteins see Anthony, 1986, 2000, 2004; Goodwin & Anthony, 1998;  Anthony & Ghosh, 1998; and www.chris-anthony.co.uk).

Top of Methanol oxidation      References for Methanol oxidation      
Illustrations for oxidation of methane, methanol and formaldehyde
TOP of Page
Electron transport and energy transduction  TOP Pages

David O’Keefe and Sandy Cross in 1980 showed that Methylobacterium extorquens and Methylophilus methylotrophus have two types of cytochrome c, named after their high and low isoelectric points – cytochrome cH and cytochrome cL (O’Keeffe & Anthony, 1980; Cross & Anthony, 1980), and this is a notable feature of all methylotrophs (Anthony, 1992). We now know that cytochrome cL is a completely unique cytochrome with no sequence identity whatsoever with any other cytochrome c (except for the CXXCH haem-binding motif). Remarkably, however, its X-ray structure shows a strong similarity to other cytochromes c (Williams et al., 2006). This novel cytochrome c is the physiological electron acceptor for MDH (Duine et al., 1979; Beardmore-Gray et al., 1983; Anthony 1992). In an investigation of proton translocation during operation of the electron transport chain for methanol oxidation we showed that this must be occurring in the periplasm (O’Keeffe & Anthony (1978), a conclusion supported by a direct demonstration of the periplasmic location of MDH by Alefounder & Ferguson (1983). The cytochrome cL is oxidised by a typical small cytochrome c (in these bacteria called cytochrome cH) which is the substrate for the oxidase, cytochrome aa3. This remarkable periplasmic electron transport chain leads to the release of 2 protons on the outside and consumption of 2 protons at the internal active site of the oxidase, leading to a protonmotive force sufficient to produce 0.7 ATP (O’Keeffe & Anthony,1978; Anthony, 1992). There has been relatively little work on the electron transport chain of methanotrophs but later studies of their genomes indicates that these are essentially the same as in Methylobacterium extorquens.

Top of Electron transport       References for Electron transport      
Illustrations for Cytochromes and Electron transport

TOP of Page
Genetics                                                            TOP of Pages   

‘Facultative methanotrophs’; genetic analysis

This topic is included to facilitate the following brief account of the history of genetics in methanotrophs, most of which are not able to grow on multicarbon compounds; that is, they are obligate methanotrophs, which makes work on mutant isolation and genetics more difficult. The isolation of Methylobacterium organophilum appeared to offer some hope, because it was first described as a facultative methanotroph and some preliminary work was done using genetic transformation to show linkage of C1-related genes (O’Connor & Hanson, 1977, 1978); it should be noted that none of this work depended on growth on methane, all experiments being done on methanol media. It is quite possible that this was not a true facultative methanotroph. Theisen & Murrell (2005) nicely summarised the situation: “Claims for the existence of facultative methanotrophs have a long and somewhat checkered history dating back almost 40 years, when Patt and colleagues first isolated and later described Methylobacterium organophilum, which was able to grow on methane or glucose (Patt et al., 1974, 1976). This was followed by other reports of facultative methanotrophs, notably strain R6, Methylobacterium ethanolicum, and Methylomonas sp. strain 761M (Lynch et al., 1980; Patel et al., 1978; Zhao et al., 1984). In most cases, either the methane-oxidizing capacity was lost or the results were not substantiated in other laboratories. For example, for M. organophilum it was hypothesized that methane oxidation genes were plasmid encoded and that the loss of growth on methane was attributable to loss of a plasmid. In the case of M. ethanolicum, the culture was shown to be a very tight syntrophic association between a methanotroph and a Xanthobacter species (Lidstrom-O’Connor et al., 1983)”.

More recently, bona fide facultative methanotrophs have been described, together with the meticulous techniques required to confirm their special status. The general problems associated with the study of facultative methanotrophs have been usefully reviewed by Theisen & Murrell (2005) and Semrau et al., (2011).

Genetics of obligate methanotrophs. 

Earlier studies of obligate methanotrophs proved them to be refractory to classical genetic analysis (Williams and Bainbridge 1971; Harwood et al. 1972; Williams et al. 1977). Although methods for mutagenesis of methane oxidation systems and for conjugal transfer of broad host range plasmids (R300B and pULB113) were successfully applied to Methylomonas albus and Methylosinus trichosporium (A1-Taho and Warner 1987; Nicolaidis and Sargent 1987; McPheat et al. 1987a, b), these were not further exploited at that time.

One of the first systems to prove amenable to genetic analysis in the methanotrophs was the nitrogen fixation system. A Klebsiella pneumoniae DNA fragment containing the nif structural genes was used to identify a clone from a Methylosinus 6 genomic bank that contained the nifD gene (Toukdarian & Lidstrom, 1984a), the identity of this gene being confirmed by construction of a chromosomal Tn5 mutant by homologous recombination. This mutant contained no detectable nitrogenase activity and was missing the nifD polypeptide on two-dimensional polyacrylamide gels (Toukdarian & Lidstrom,

The nitrogen fixation genes were similarly some of the first to be involved in genetic analysis by Murrell and colleagues in the University of Warwick, UK (Oakley & Murrell, 1988) who used the nifH gene from Klebsiella pneumoniae, which codes for the Fe protein component of nitrogenase, as a probe to detect nifH homologues in total cellular DNA from 13 obligate methane oxidizing bacteria. All but one of those strains that had previously been shown capable of fixing dinitrogen contained sequences homologous to nifH.

Two key papers in the further development of methylotroph genetics were published in 1986 by David Nunn and Mary Lidstrom (at the University of Washington, Seattle, USA), who reported the genetic analysis of functions necessary for methanol oxidation (Mox functions) in the facultative methanol-utilizer Methylobacterium AM1, which suggested that at least ten different gene products are involved (Nunn & Lidstrom, 1986a,b). The ten mox genes included two encoding the methanol dehydrogenase MDH) and its specific electron acceptor cytochrome cL, and eight were involved in assembly and regulation. The genetics of the assimilation pathway was initiated in this same organism (Stone & Goodwin, 1989) and transposon mutagenesis of methanol oxidation genes were described (Lee et al., 1991).  Lidstrom’s work on the methanol-utiliser Methylobacterium AM1was then extended by her group to the obligate methanotrophs, using a portion of the Methylobacterium AM1 MDH structural gene (moxF) as a hybridization probe to isolate and characterize the moxF genes from two obligate methanotrophs, Methylococcus capsulatus and Methylomonas alba (Stephens et al., 1988). This work on the methanol oxidation system in methanotrophs was later extended to investigate the genetics of methanol oxidation in Methylomonas sp. A4 (Waechter-Brulla et al., 1993) and to identify the promoter for expression of the gene for methanol dehydrogenase (moxF) in the Type I methanotroph, Methylobacter albus BG (Chistoserdova et al., 1994).

At about the time that Mary Lidstrom’s group started their investigations of the genetics of the methanol oxidation system, work started in the University of Warwick (UK) in the groups of Murrell and Dalton on the genetics of methanotrophs, especially in relation to the methane monooxygenase (MMO) (Mullens & Dalton, 1987; McPheat et al., 1987a,b). In this work, oligonucleotide probes specific to the N-terminal amino acid sequence of individual polypeptides of the soluble MMO from MethyIococcus capsuIatus (Bath) were used to identify MMO genes. Subsequently, the genes encoding the β and γ subunits of MMO were shown to be linked in this organism, and analysis of a recombinant plasmid containing 12 kilobases of Methylococcus DNA provided the first evidence for the localization and linkage of genes encoding the methane monooxygenase enzyme complex, DNA sequence analysis providing the complete sequence of the genes encoding the β and γ subunits of MMO (Stainthorpe et al., 1989). These studies were extended and expanded in Murrell’s group, in collaboration with Lidstrom’s group, mainly concentrating on the genetics of the two MMOs and their regulation (Stainthorpe et al., 1990; Cardy et al., 1991a,b; Semrau et al., 1995; Murrell, 1992, 1994). The year 1994 is the point in time when ‘History’ becomes ‘Current affairs’.

Top of Genetics         References for Genetics         TOP of Page
REFERENCES   These are arranged in groups corresponding to the section headings

Introduction and carbon assimilation;       Methane oxidation;
Methanol oxidation;
      Electron transport;      Genetics

Introduction and Carbon assimilation      Back to text

Anthony (1982). The Biochemistry of Methylotrophs. Academic Press, London.

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.

Brown, L.R., Strawinski, R.J. and McCleskey, C.S. (1964). The isolation and characterization of
Methanomonas methanooxidans. Canadian Journal of Microbiology10, 791-799.

Dworkin, M. and Foster, J. W. (1956). Studies on Pseudomonas methanica (Sohngen) nov. comb. J. Bacteriol.72, 646-659.

Ferenci, T., Strom, T. and Quayle, J. R. (1974). Purification and properties of 3-hexulose phosphate synthase and phospho-3-hexuloisomerase from Methylococcus capsulatus.  Biochem. J. 144, 477-486.

Foster, J.W. and Davis, R. H. (1966). A methane-dependent coccus, with notes on classification
and nomenclature of obligate, methane-utilizing bacteria. J. Bacteriol.91, I924-1931.

Johnson, P. A. and Quayle, J. R. (1965). Microbial Growth on C1 Compounds.
Synthesis of cell constituents by methane- and methanol-grown Pseudomonas methanica Biochem . J.95, 859-867.
Kaneda, T. and Roxburgh, J. M. (1959a). A methanol-utilizing bacterium: II. Studies on the pathway of methanol assimilation. Canad. J. Microbiol.  5, 187-195.

Kaneda, T. and Roxburgh, J. M. (1959b). Serine as an intermediate in the assimilation of methanol by a Pseudomonas. Biochim. biophys.Acta, 33, 106-110.

Kemp, M. B. and Quayle, J. R. (1965). Incorporation of C1 units into allulose phosphate by methane-grown Pseudomonas methanica . Biochim. Biophys. Acta 107, 174-176.
Kemp, M. B. and Quayle, J. R. (1966).  Microbial growth on C1 compounds. Incorporation of C1 units into allulose phosphates by extracts of Pseudomonas methanica. Biochem. J. 99, 41 - 48.

Kemp, M.B. and Quayle, J.R. (1967). Uptake of [14C]formaldehyde and [14C]formate by methane-grown Pseudornonas methanica and determination of the hexose labelling pattern after brief incubation with [14C]methanol. Biochem. J.102, 94-102.

Kemp, M. B. (1974). Hexose phosphate synthase from Methylococcus capsulatus makes           D-arabino-3-hexulose phosphate.  Biochem. J. 139, 129-134.

Large, P.J., Peel, D. and Quayle, J.R. (1961).  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.

Large, P.J. Peel, D. and Quayle, J.R. (1962a) 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.

Large, P.J. Peel, D. and Quayle, J.R. (1962b).  Microbial growth on C1 compounds: 4. Carboxylation of phosphoenolpyruvate in methanol-grown Pseudomonas AM1. Biochem. J. 85, 243-250.

Large, P.J. and  Quayle, J.R. (1963).  Microbial growth on C1 compounds: 5. Enzyme activities in extracts of Pseudomonas AM1. Biochem. J., 87 386-396.

Leadbetter, E.R. and Foster, J.W. (I958).Studies on some methane utilizing bacteria. Archiv. Fur Mikrobiologie 30, 91-118.

Lawrence, A.J and Quayle, J.R. (1970). Alternative carbon assimilation pathways in
methane-utilizing bacteria.  Journal of General Microbiology 63, 371-374.

Lawrence, A.J., Kemp, M.B. and Quayle, J.R. (1970). Synthesis of cell constituents by methane-grown Methylococcus capsulatus and Methanomonas methanooxidans. Biochem. J.116, 631-639.

Orla-Jensen, S.  (1909). Die Hauptlinien des Naturlichen Bakteriensystems. Centr. Bakt.
Parasitenk., Abt. II, 22, 305-346.

Peel, D. and Quayle, J.R. (1961). Microbial growth on C1 compounds. 1. Isolation and characterization of Pseudomonas AM!. Biochem. J. 81, 465-469.

Quayle, J.R., Fuller, R.C., Benson, A.A. and Calvin, M. (1954). Enzymatic carboxylation of ribulose diphosphate. J. Amer. Chem. Soc. 76, 3610-3611.

Quayle, J. R. and Keech, D. B. (1959a). Carbon assimilation by Pseudomonas oxalaticus (OX 1). 1. Formate and carbon dioxide utilization during growth on formate. Biochem. J. 72, 623-630.

Quayle, J. R. and Keech, D. B. (1959b). Carbon assimilation by Pseudomonas oxalaticus (OX 1). 2. Formate and carbon dioxide utilization by cell-free extracts of the organism grown on formate. Biochem. J. 72, 631-637.

Salem, A.R., Hacking , A.J. and Quayle, J.R. (1973). Cleavage of malyl-coenzyme A into acetyl-coenzyme A and glyoxylate by Pseudomonas AM1 and other Cl-unit-utlizing bacteria. Biochem. J. 136, 89-96.

Sohngen, N. L. (1906). Zentbl. Bakt. ParasitKde. (Abt. II) 15, 513.

Sohngen, N. L (1910).  Sur le role du methane dans la vie organique. Rec. trav. chim.,
29, 238-274.

Strom, T., Ferenci, T. and Quayle, J. R. (1974). The carbon assimilation pathways of Methylococcus capsulatus, Pseudomonas methanica and Methylosinus trichosporium (Ob3b) during growth on methane.  Biochem. J. 144, 465-476.

Whittenbury, R., Phillips, K.C. and Wilkinson, J.F.  (1970a). Enrichment, Isolation and some properties of methane-utilizing bacteria. Journal of General Microbiology61, 205-218.

Whittenbury, R., Stephanie L. Davies, S.L. and Davey, J.F. (1970b). Exospores and cysts formed by methane-utilizing bacteria.Journal of General Microbiology61, 219-226.  

Methane oxidation              Back to text

Anthony, C. (2008). A tribute to Howard Dalton and methane monooxygenase.  Science Progress 91, 401–415.

Colby, J., Dalton, H. and Whittenbury, R. (1975). An improved assay for bacterial methane monooxygenase: some properties of the enzyme from Methylomonas methanica. Biochem. J. 151, 459-462.

Colby, J., Stirling, D. and Dalton, H. (1977) The soluble methane monooxygenase of Methylococcus capsulatus (Bath): its ability to oxygenate n-alkanes, n-alkenes, ethers, and acyclic, aromatic and heterocyclic compounds. Biochem. J. 165, 395-402.

Colby, J. and Dalton, H. (1978). Resolution of the methane monooxygenase of Methylococcus capsulatus (Bath) into three components. Biochem. J. 171, 461-468.

Colby, J. and Dalton, H. (1979). Characterisation of the second prosthetic group of the flavoenzyme NADH-acceptor reductase (component C) of the methane monooxygenase of Methylococcus capsulatus( Bath). Biochem.J . 177, 903-908.

Dalton, H. (2005).  The Leeuwenhoek Lecture 2000: The Natural and Unnatural History of Methane-Oxidizing Bacteria.  Philosophical Transactions: Biological Sciences, 360, 1207-1222.

Higgins, I.J. and Quayle, J.R. (1970). Oxygenation of methane by methane-grown Pseudomonas methanica and Methanomonas methanooxidans. Biochem. J. 118, 201 -2018.

Leadbetter, E.R. and Foster, J.W. (1959). Incorporation of molecular oxygen in bacterial cells utilizing hydrocarbons for growth. Nature 184, 1428-1429.

Ribbons, D. W. and Michalover, J. L. (1970).  Methane oxidation by cell-free extracts of Methylococcus capsulatus.  FEBS Lett. 11, 41-44.

Ribbons, D. W. (1975). Oxidation of C1 compounds by particulate fractions from Methylococcus capsulatus: distribution and properties of methane-dependent reduced nicotinamide adenine dinucleotide oxidase (methane hydroxylase). J. Bacteriol.122, 1351-1363.

Stanley, S. H., Prior, S.D., Leak, D.J. and Dalton, H. (1983). Copper stress underlies the fundamental change in intracellular location of methane monooxygenase in methane-oxidising organisms: studies in batch and continuous cultures. Biotechnol. Lett. 5, 487-492.

Stirling, D.I. and Dalton, H. (1979). A comparison of the substrate and electron-donor specificities of the methane mono-oxygenases from three strains of methane-oxidizing bacteria.
Biochem. J. (1979) 177, 361-364.

Tonge, G. M., Harrison, D. E. F. and Higgins, I. J. (1977). Purification and properties of the methane monooxygenase system from Methylosinus trichosporium OB3b. Biochem. J. 161, 333-344.

Methanol oxidation            Back to text

Anthony, C (1986). The bacterial oxidation of methane and methanol. Adv. Microbial. Physiol.  27, 113–210.

Anthony, C. (2000). Methanol dehydrogenase, a PQQ-containing quinoprotein dehydrogenase. Subcell. Biochem. 35, 73–118.

Anthony, C. (2004). The quinoprotein dehydrogenases for methanol and glucose. Archives of Biochemistry and Biophysics 428, 2–9.

Anthony, C. and Zatman, L.J. (1964a). The methanol-oxidizing enzyme of Pseudomonas sp. M27. Biochem. J. 92, 614-621.

Anthony, C. and Zatman, L.J. (1964b). Isolation and properties of Pseudomonas sp. M27. Biochem. J. 92, 609-614.

Anthony, C. and Zatman, L.J. (1965). The alcohol dehydrogenase of Pseudomonas sp. M27. Biochem. J. 96, 808-812.

Anthony, C. and Zatman, L.J. (1967a). The microbial oxidation of methanol: purification and properties of the alcohol dehydrogenase of Pseudomonas sp. M27. Biochem. J. 104, 953-959.

Anthony, C. and Zatman, L.J. (1967b). The microbial oxidation of methanol: The prosthetic group of alcohol dehydrogenase of Pseudomonas sp. M27; A new oxidoreductase prosthetic group. Biochem. J. 104, 960-969.

Chan, H.T.C. and Anthony, C. (1992). The mechanism of inhibition by EDTA and EGTA of methanol oxidation by methylotrophic bacteria. FEMS Microbiol Lett 96, 231-234.

Goodwin, P.M. and Anthony, C. (1988). The biochemistry, physiology and genetics of PQQ and PQQ-containing enzymes. Advances in Microbial Physiology, 40, 1-80.

Harrington, A. A. and Kallio, R. E. (1960). Oxidation of methanol and formaldehyde by Pseudomonas methanica. Canadian Journal of Microbiology 6, 1-7.

Hauge, J.G. (1964). Glucose dehydrogenase of Bacterium anitratum: An enzyme with a novel prosthetic group. J. Biol. Chem. 239, 3630—3639.

Johnson, P.A. and Quayle, J.R. (1964). Microbial Growth on C1 Compounds. 6. Oxidation of methanol, formaldehyde and formate by methanol-grown Pseudomonas AM1.  Biochem. J. 93, 281-290.

Kaneda, T. and Roxburgh, J. M. (1959a). A methanol-utilizing bacterium: II. Studies on the pathway of methanol assimilation. Canad. J. Microbiol.  5, 187-195.

Nunn, D.N., Day, D.J. and Anthony, C. (1989). The second subunit of methanol dehydrogenase of Methylobacterium extorquens AM1. Biochem. J. 260, 857-862.

Patel, R. N. and Felix, A. (1976). Microbial oxidation of methane and methanol: crystallization and properties of methanol dehydrogenase from Methylosinus sporium. J. Bacteriol.128, 413-424.

Patel, R. N., Bose, H.R., Mandy, W.J. and Hoare, D.S. (1972). Physiological studies of methane and methanol-oxidizing bacteria: comparison of a primary alcohol dehydrogenase from Methylococcus capsulatus (Texas strain) and Pseudomonas species M27. J. Bacteriol.
110, 570-577.

Patel, R.N., Mandy, W.J. and Hoare, D.S. (1973). Physiological studies of methane- and methanol-oxidizing bacteria: immunological comparison of a primary alcohol dehydrogenase from Methylococcus capsulatus and Pseudomonas sp. M27. J. Bacteriol. 113, 937-945.

Salisbury, S.A., Forrest, H.S., Cruse, W.B.T. and Kennard, O. (1979). A novel coenzyme from bacterial primary alcohol dehydrogenases. Nature 280, 843–844.

Wadzinski, A. M., and Ribbons, W.D. (1975). Oxidation of Cl compounds by particulate fractions from Methylococcus capsulatus: properties of methanol oxidase and methanol dehydrogenase. J. Bacteriol. 122, 1364-1374.

Williams, P.A.,  Coates, L., Mohammed, F., Gill, R.,  Erskine, P.T., Coker, A.,  Wood, S.P., Anthony, C. and Cooper, J.B. (2005). The atomic resolution structure of methanol dehydrogenase from Methylobacterium extorquens. Acta Crystallogr D Biol Crystallogr. 61, 75-79.

Electron transport          Back to text

Alefounder, R. and Ferguson, S. J . (1981). A periplasmic location for methanol dehydrogenase
from Paracoccus denitrificans; implications for proton pumping by cytochrome aa3. Biochemical and Biophysical Research Communications98, 778-784.

Anthony, C. (1992). The c-type cytochromes of methylotrophic bacteria. Biochim. Biophys. Acta1099, 1-15.

Beardmore-Gray, M., O'Keeffe, D.T. and Anthony, C. (1983). The methanol:cytochrome c oxidoreductase activity of methylotrophs. Journal of General Microbiology 129, 923-933.
Cross, A.R. and Anthony, C. (1980). The purification and properties of the soluble cytochromes c of the obligate methylotroph Methylophilus methylotrophus. Biochem. J. 192, 421-427.

Duine, J.A., Frank, J. and Ruiter, L.G. (1979). Isolation of a methanol dehydrogenase with a
functional coupling to cytochrome c. J. Gen. Microbiol. 115, 523-526.

O'Keeffe, D.T. and Anthony, C. (1978). The microbial metabolism of C1 compounds. The stoichiometry of respiration-driven proton translocation in Pseudomonas AM1 and in a mutant lacking cytochrome c. Biochem. J. 170, 561-567.

O'Keeffe, D.T. and Anthony, C. (1980). The two cytochromes c in the facultative methylotroph Pseudomonas  AM1. Biochem. J. 192, 411-419.

Williams, P., Coates, L., Mohammed, F., Gill, R., Erskine, P., Bourgeois, D., Wood, S.P.,  Anthony, C. and Cooper, J.B. (2006). The 1.6 A° X-ray structure of the unusual c-type cytochrome, cytochrome cL, from the methylotrophic bacterium Methylobacterium extorquens. J. Mol. Biol. 357, 151–162.

Genetics                  Back to text

AI-Taho, N.M. and Warner, P.J. (1987). Restoration of phenotype in Escherichia coli auxotrophs by pULBll3-mediated mobilization from methylotrophic bacteria. FEMS Microbiol. Lett 43, 235- 239.

Cardy, D. L. N., Laidler, V., Salmond, G. P. C. and Murrell, J. C. (1 991a). Molecular analysis of the methane monooxygenase (MMO) gene cluster of Methylosinus trichosporium OB3b. Mol. Microbiol. 5, 335-342.

Cardy, D. L. N., Laidler, V., Salmond, G. P. C. and Murrell, J. C. (1991 b). The methane monooxygenase gene cluster of Metbylosinus trichosporium : cloning and sequencing of the mmoC gene. Arch. Microbiol. 56, 477-483.

Chistoserdova, L., Kuhn, M and Lidstrom M.E. (1994). Identification of a promoter region for mxaF (moxF) from the Type I methanotroph, Methylobacter albus BG8. FEMS Microbiol. Lett. 121, 343-348.

Harwood, J.J., Williams, E.and Bainbridge, B.W. (1972). Mutation of the methane oxidizing bacterium Methylococcus capsulatus. J. Appl. Bacteriol. 35, 99-108.

Lee, K. E., Stone, S., Goodwin, P. M. and Holloway, B. W. (1991). Characterization of transposon insertion mutants of Metbylobacterium extorquens AM1 (Metbylobacterium strain AM1) which are defective in methanol oxidation. J Gen Microbiol. 37, 895-904.

Lidstrom-O’Connor, M.E., Fulton, G.L. and Wopat, A.E. (1983) ‘Methylobacterium ethanolicum’: a syntrophic association of two methylotrophic bacteria. J. Gen. Microbiol. 129, 3139–3148.

Lynch, M. J., Wopat, A.E. and O’Connor, M.L. (1980).  Characterization of two new facultative methanotrophs. Appl. Environ. Microbiol. 40, 400–407.

McPheat, W.L., Mann, N.H. and Dalton, H. (1987a). Transfer of broad host range plasmids to the type 1 obligate methanotroph Methylomonas albus. FEMS Microbiol. Lett. 41, 185-188.

McPheat, W.L., Mann, N.H. and Dalton, H. (1987b). Isolation of mutants of the obligate methanotroph Methylococcus albus defective in growth on methane. Arch. Microbiol. 148, 40-43.

Mullens, I. A . and Dalton, H. (1987). Cloning of the gamma subunit of methane monooxygenase from Methylococcus capsulatus. Bio-Technology 5, 490—93.

Murrell, J. C. (1992). Genetics and molecular biology of methanotrophs. FEMS Microbiol. Rev. 88, 233-248.

Murrell, J. C. (1994). Molecular genetics of methane oxidation. Biodegradation 5, 145-159.

Nicolaidis, A.A. and Sargent, A.W. (1987). Isolation of methane monooxygenase deficient mutants from Methylosinus trichosporium OB3b using dichloromethane. FEMS Microbiol. Lett. 41, 47- 52.

Nunn, D.N. and Lidstrom, M.E. (1986a). Isolation and complementation analysis of 10 methanol oxidation mutant classes and identification of the methanol dehydrogenase structural gene of Methylobacterium sp. Strain AMI.  J. Bacteriol. 166, 581-590.

Nunn, D.N. and Lidstrom, M.E. (1986b). phenotypic characterization of 10 methanol oxidation mutant
classes in Methylobacterium sp. Strain AM1. J. Bacteriol. 166, 591-597.

Oakley, C . and Murrell, J. C. (1988). NifH genes in the obligate methane oxidizing bacteria. FEMS Microbiol. Lett. 49, 53- 58.

O'Connor, M. L. , Wopat, A . , Hanson, R. S. (1977). Genetic transformation in Methylobacterium organophilum. J. Gen. Microbiol. 98,265-272.

O 'Connor, M. L.  and Hanson, R. S. (1978). Linkage relationships between mutants of Methylobacterium organophilum impaired in their ability to grow on one-carbon compounds. J. Gen. Microbiol. 104, 105-111.

Patel, R. N., Hou, C.T. and Felix, A.. 1978. Microbial oxidation of methane and methanol: isolation of methane-utilizing bacteria and characterization of a facultative methane-utilizing isolate. J. Bacteriol. 136, 352–358.

Patt, T. E., Cole, G. C., Bland, J. and Hanson, R. S. (1974). Isolation and characterization of bacteria that grow on methane and organic compounds as sole sources of carbon and energy. Journal of Bacteriology 120, 955-964.

Patt,T . E., Cole, G. C. and Hanson, R. S. (1976). Methylobacterium, a new genus of facultatively
methylotrophic bacteria. International Journal of Systematic Bacteriology 26, 226-229.

Semrau, J.D., Chistoserdov, A, Lebron, J., Costello, A., Davagnino, J.,. Kenna, E., Holmes, A.J.,  Finch, R., Murrell, J.C. and Lidstrom. M.E. (1995). Particulate methane monooxygenase genes in methanotrophs. J. Bacteriol. 177, 3071-3079.

Semrau, J.D., DiSpirito, A.A. and Vuilleumier, S. (2011). Facultative methanotrophy: false leads, true results, and suggestions for future research. FEMS Microbiol. Lett. 323, 1–12.

Stainthorpe, A. C., Murrell, J. C.,  Salmond, G. P. C., Dalton, H. and Lees, V. (1 989). Molecular analysis of methane monooxygenase  from Methylococcus capsulatus (Bath). Arch. Microbiol. 152, 154-159.

Stainthorpe, A. C., Lees, V., Salmond, G. P. C., Dalton, H. and Murrell, J. C. (1990). The methane monooxygenase gene cluster of Methylococcus capsulatus (Bath). Gene 91, 27-34.

Stephens, R. L ., Haywood, M. G. and Lidstrom, M. E. (1988). Identification of putative methanol dehydrogenase (moxF) structural genes in methylotrophs and cloning of moxF genes from
Methylococcus capsulatus Bath and Methylomonas albus BG8. J. Bacteriol. 170, 2063—69.

Stone, S. and Goodwin, P.M. (1989). Characterization and complementation of mutants of Methylobacterium extorquens AM1 which are defective in C1 assimilation. J. Gen. Microbiol. 135, 227 – 235.

Facultative Methanotrophs Revisited. J. Bact. 187, 4303–4305.

Toukdarian, A. E .  and Lidstrom, M. E.  (1984a). Molecular construction and characterization of nil mutants of the obligate methanotroph, Methylosinus sp. strain 6.  J. Bacteriol. 157, 979-983.

Toukdarian, A. E .  and Lidstrom, M. E.  (1984b). DNA hybridization analysis of the nif region of Methylosinus 6.  J. Bacteriol. 57,  925-930.

Waechter-Brulla, D., DiSpirito, A.A. and M.E. Lidstrom, M.E. (1993). Genetics of methanol oxidation in Methylomonas sp. A4. J. Bacteriol. 175, 3767-3775.

Williams, E. and Bainbridge, B.W. (1971). Genetic transformation in Methylococcus capsulatus. J. Appl. Bacteriol. 34, 683 – 687. 

Williams, E., Shimmin, M.A. and Bainbridge, B.W. (1977). Mutation in the  obligate methylotrophs Methylococcus capsulatus and Methylomonas albus. FEMS Microbiol. Lett. 2, 293 – 296.

Zhao, S. J., and Hanson, R.S. (1984). Variants of the obligate methanotroph isolate 761M capable of growth on glucose in the absence of methane. Appl. Environ. Microbiol. 48, 807–812.

TOP of References      Top of Page










logo design