Mutants of Escherichia coli producing pyrroloquinoline quinone

Biville, Francis; Turlin, Évelyne; Gasser, F.

doi:10.1099/00221287-137-8-1775

Cited by 26 publications

(21 citation statements)

References 19 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…E. coli synthesizes an inactive glucose dehydrogenase apoprotein that can be activated with PQQ (18), and it is possible that some strains contain genes required for parts of the PQQ biosynthetic pathway. In at least one strain, mutants that are now able to produce PQQ can be isolated (7). Further work is under way to determine the functions of each of these genes involved in PQQ biosynthesis in M. extorquens AML.…”

Section: Discussionmentioning

confidence: 99%

Isolation, phenotypic characterization, and complementation analysis of mutants of Methylobacterium extorquens AM1 unable to synthesize pyrroloquinoline quinone and sequences of pqqD, pqqG, and pqqC

et al. 1994

View full text Add to dashboard Cite

Aerobic gram-negative methylotrophs oxidize methanol to formaldehyde by using a methanol dehydrogenase that has pyrroloquinoline quinone (PQQ) as a prosthetic group. Seventy-two mutants which are unable to grow on methanol unless the growth medium is supplemented with PQQ have been isolated in the facultative methanol utilizer Methylobacterium extorquens AM1. In addition, 12 previously isolated methanol oxidation mutants of M. extorquens AM1 were shown to be able to grow on methanol in the presence of PQQ. These putative PQQ biosynthesis mutants have been complemented by using previously isolated clones containing M. extorquens AM1 DNA, which were known to contain genes necessary for oxidation of methanol to formaldehyde (mox genes (27). MDH is a tetrameric enzyme located in the periplasm that contains pyrroloquinoline quinone (PQQ) as the prosthetic group and also contains Ca2+ (37,40). PQQ was first identified as the prosthetic group of MDH and is now also known to be the prosthetic group of a few other bacterial dehydrogenases that oxidize alcohols or sugars (9). The biosynthetic pathway of PQQ has not yet been determined, but the biosynthetic precursors are known to be tyrosine and glutamate (19).

show abstract

Section: Discussionmentioning

confidence: 99%

Isolation, phenotypic characterization, and complementation analysis of mutants of Methylobacterium extorquens AM1 unable to synthesize pyrroloquinoline quinone and sequences of pqqD, pqqG, and pqqC

et al. 1994

View full text Add to dashboard Cite

show abstract

“…The following E. coli strains were used : FB8 (prototrophic E. coli K-12 strain) ; Aptsgal [A(ptsHZ-crr) : : Km galP : : TnlO, derived from FB8 and renamed PPA3221; EF260 (Glc+, derived from Aptsgal) (isolated and described by Biville et al, 1991); PPA533 (gcd::Cm, derived from EF260); and CT690 (recB22 recC22 thi-1 thr-1 leu-6 lacy1 mtl-2 xyl-1 ara-14 galK2 his-4 proA2 arg-63 rpsL31 tsx-33 sup-37 sbcB15, from Dr M. Tsuda, Okayama University, Japan). The pre-culture was washed twice with 10mM potassium phosphate buffer (pH 7.5) and resuspended in the same buffer at the same concentration as the original pre-culture.…”

Section: Methodsmentioning

confidence: 99%

Escherichia coli is unable to produce pyrroloquinoline quinone (PQQ)

et al. 1997

View full text Add to dashboard Cite

Many bacteria can synthesize the cofactor pyrroloquinoline quinone (PQQ), a cofactor of several dehydrogenases, including glucose dehydrogenase (GCD). Among the enteric bacteria, Klebsiella pneumoniae has been shown to contain the genes required for PQQ biosynthesis. Escherichia coli and Salmonella typhimurium were thought to be unable to synthesize PQQ but it has been reported that strain EF260, a derivative of E. coli FB8, can synthesize PQQ after mutation and can oxidize glucose to gluconate via the GCD/PQQ pathway (F. Biville, E. Turlin & F. Gasser, 1991, J Gen Micmbioll37,1775-1782). We have reinvestigated this claim and conclude that it is most likely erroneous. (i) Strain EF260, isolated originally by Biville and coworkers, was unable to synthesize a holo-enzyme GCD unless PQQ was supplied to the growth medium. No GCD activity could be detected in membrane fractions. (ii) The amount of PQQ detected in the growth medium of EF260 was very low and not very different from that found in a medium with its parent strain or in a medium containing no cells. (iii) EF260 cells were unable to produce gluconate from glucose via the PQQ/GCD pathway. (iv) Introduction of a gcd::Cm deletion in EF260, eliminating GCD, did not affect glucose metabolism. This suggested a pathway for glucose metabolism other than the PQQ/GCD pathway. (v) Glucose uptake and metabolism in EF260 involved a low-affinity transport system of unknown identity, followed most likely by phosphorylation via glucokinase. It is concluded that E. coli cannot synthesize PQQ and that it lacks genes required for PQQ biosynthesis.

show abstract

“…E. coli K-12 derivatives are capable of synthesizing the apo-glucose dehydrogenase (apo-GDH) but not the cofactor, pyrroloquinoline quinone (PQQ), which is essential for the formation of the holoenzyme (3,5). When PQQ is present in the medium, the holoenzyme is reconstituted and then E. coli is capable of oxidizing glucose to gluconate (7).…”

mentioning

confidence: 99%

Cloning and expression of a gene cluster encoding three subunits of membrane-bound gluconate dehydrogenase from Erwinia cypripedii ATCC 29267 in Escherichia coli

1997

View full text Add to dashboard Cite

We have cloned the gene cluster encoding three subunits of membrane-bound gluconate dehydrogenase (GADH) from Erwinia cypripedii ATCC 29267 in Escherichia coli by performing a direct-expression assay. The positive clone converted D-gluconate to 2-keto-D-gluconate (2KDG) in the culture medium. Nucleotide sequence analysis of the GADH clone revealed that the cloned fragment contained the complete structural genes for a 68-kDa dehydrogenase subunit, a 47-kDa cytochrome c subunit, and a 24-kDa subunit of unknown function and that the genes were clustered with the same transcriptional polarity. Comparison of the deduced amino acid sequences and the NH 2 -terminal sequences determined for the purified protein indicated that the dehydrogenase, cytochrome c, and 24-kDa subunits contained typical signal peptides of 22, 19, and 42 amino acids, respectively. The molecular masses of the processed subunits deduced from the nucleotide sequences (65, 45, and 20 kDa) coincided well with the molecular masses of subunits estimated by sodium dodecyl sulfatepolyacrylamide gel electrophoresis. In E. cypripedii and recombinant E. coli, the GADH was constitutively formed and the activity of GADH was enhanced more than twofold by addition of D-gluconate to the medium. The conversion of D-glucose to D-gluconate, 2-keto-D-gluconate (2KDG), and 2,5-diketo-D-gluconate (25DKG) in several species of the genera Acetobacter, Gluconobacter, and Erwinia is mediated by membrane-bound dehydrogenases linked to the cytochrome chains located in the cytoplasmic membrane of the bacterium (1, 31). Membrane-bound gluconate dehydrogenases (GADHs), which catalyze the production of 2KDG from D-gluconate, have been purified and characterized from Pseudomonas aeruginosa (16,19), P. fluorescens (17), Klebsiella pneumoniae (17), Serratia marcescens (17), and Gluconobacter dioxyacetonicus (28). The subunit structures and enzymatic properties of these GADHs were found to be very similar (17). These GADHs consisted of three subunits; namely, flavoprotein, cytochrome c, and a third subunit whose function is unknown. Despite these findings, no further work on gene structures has been reported.In the present work, we describe the direct-expression cloning of the gene cluster encoding three subunits of membranebound GADH from Erwinia cypripedii ATCC 29267. E. coli K-12 derivatives are capable of synthesizing the apo-glucose dehydrogenase (apo-GDH) but not the cofactor, pyrroloquinoline quinone (PQQ), which is essential for the formation of the holoenzyme (3, 5). When PQQ is present in the medium, the holoenzyme is reconstituted and then E. coli is capable of oxidizing glucose to gluconate (7). It has also been reported that the expression of PQQ synthase genes in E. coli resulted in GDH activity in the absence of exogenous PQQ (13). Therefore, we tried to convert D-glucose to 2KDG via D-gluconate by using recombinant E. coli harboring the cloned GADH gene in the presence of PQQ. MATERIALS AND METHODSBacterial strains, media, and culture conditions. E. cypripedii ATCC ...

show abstract

Mutants of Escherichia coli producing pyrroloquinoline quinone

Cited by 26 publications

References 19 publications

Isolation, phenotypic characterization, and complementation analysis of mutants of Methylobacterium extorquens AM1 unable to synthesize pyrroloquinoline quinone and sequences of pqqD, pqqG, and pqqC

Isolation, phenotypic characterization, and complementation analysis of mutants of Methylobacterium extorquens AM1 unable to synthesize pyrroloquinoline quinone and sequences of pqqD, pqqG, and pqqC

Escherichia coli is unable to produce pyrroloquinoline quinone (PQQ)

Cloning and expression of a gene cluster encoding three subunits of membrane-bound gluconate dehydrogenase from Erwinia cypripedii ATCC 29267 in Escherichia coli

Contact Info

Product

Resources

About