Catabolism of pteridine cofactors

Rembold, Heinz; Metzger, H.; Sudershan, P.; Gutensohn, Wolf

doi:10.1016/0304-4165(69)90042-7

Cited by 26 publications

(5 citation statements)

References 21 publications

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“…This picture, as expected, is not too dissimilar but is nevertheless distinguishable from the lumazine spectra, shown in Pteridines occur naturally in both eukaryotic and prokaryotic organisms, and are biosynthesized starting from purines (26)(27)(28). In most cases the terminal catabolites in pteridine degradation are simple pteridines and lumazines (29)(30)(31). Though lumazines are usually formed by catabolic pathways, at least one, namely 6,7-dimethyl-8-ribityllumazine, is involved in anabolic metabolism, riboflavin biosynthesis, in the organisms Ashbya gossypii, Eremothecium ashbyii, mutants of Bacillus subtilis, and others (7,(32)(33)(34)(35).…”

Section: Resultsmentioning

confidence: 99%

Separation and structure of the prosthetic group of the blue fluorescence protein from the bioluminescent bacterium Photobacterium phosphoreum

Koka

Lee²

1979

Proc. Natl. Acad. Sci. U.S.A.

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ABSTRACr The highly fluorescent prosthetic group of the blue fluorescence protein purified from the bioluminescent bacterium Photobacterium phosphoreum has been dissociated and separated from its apoprotein by affinity chromatography on Cibacron Blue-Sepharose. It has been identified as 6,7-dimethyl-8(1'-D-ribityl)lumazine by several methods of characterization, all of which gave results identical to those for an authentic sample. In neutral solution, absorption maxima are at 407, 275 (shoulder), and 256 nm, with a single fluorescence maximum at 491 nm. The proton magnetic resonance spectrum exhibits a singlet at 2.66 ppm corresponding to the methyl substituted at the 6 position of lumazine and a multiplet centered at 3.85 ppm corresponding to the C-2'-5' protons of the ribityl group. A Raman spectrum was obtained by the technique of coherent anti-Stokes Raman scattering and the RF values by paper chromatography were determined in four solvent systems. The isolated compound was readily transformed into riboflavin by riboflavin synthetase. Fifty grams (dry weight) of P. phosphoreum contains at least 20 mg of this lumazine derivative, an amount comparable to that found in other microorganisms classified as riboflavin overproducers. The overproduction of this lumazine in this case apparently has to do with its function in the generation of bioluminescence.A blue fluorescence protein was isolated from extracts of the bioluminescent bacterium Photobacterium phosphoreum by Gast and Lee (1); the protein was shown to meet the requirements of the in vivo bioluminescence emitter by three significant observations: (i) its fluorescence emission is identical to the bioluminescence spectrum both in position of the maximum, 476 nm, and in spectral distribution; (ii) the in vitro bioluminescence reaction of reduced flavin mononucleotide, luciferase, oxygen, and long-chain aliphatic aldehyde is stimulated by the addition of this protein both in terms of the decay rate of light emission and the total light yield; and (iii) the in vitro bioluminescence spectrum is shifted to match that of the in vivo when this protein is included in the in vitro reaction (1). The blue fluorescence protein is associated with luciferase and has a molecular weight of 22,000 (1). The protein-bound prosthetic group is highly fluorescent, with a quantum yield of 0.45 (1). Alteration of solution conditions perturbs this fluorescence to match either the in vitro or the in vivo bioluminescence of P. phosphoreum (1).The blue fluorescence protein was further purified by affinity chromatography on a Cibacron Blue-Sepharose column, which removed all traces of flavin and other fluorescent impurities (2, 3). The purified protein showed absorption maxima at 412 and 274 nm with a single absorption band in the visible region (3).The finding of a blue fluorescence protein also in another species of bioluminescent bacterium, Photobacterium fischeri (4), suggests that the fluorophore functions in some way in the bioluminescence and therefore its chemical structure sho...

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Section: Resultsmentioning

confidence: 99%

Separation and structure of the prosthetic group of the blue fluorescence protein from the bioluminescent bacterium Photobacterium phosphoreum

Koka

Lee²

1979

Proc. Natl. Acad. Sci. U.S.A.

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“…The numbers in parentheses are the estimated half-times (min) for the auto-oxidation of (6R)BH4 under the same reaction conditions. BH2 percentages are defined in Table 2 Buffer [19,201, and the isolation from rat liver of essentially pure BH2 after purification at 5°C [8]. We conclude that in phosphate buffer, at neutral pH, the tautomeric pathway leading to BH2 becomes less favored as the temperature is increased.…”

Section: -)mentioning

confidence: 98%

“…The auto-oxidation of BH, at elevated temperatures also results in a significant loss of the alkyl side-chain, i.e. boiling at pH 7.5 [19] or autoclaving at 121°C at pH 4.0 [20]. Furthermore, Armarego et al [21] demonstrated that PH2 was the major product of the autooxidation of BH4 at room temperature (25°C) in 0.09 M Tris pH 7.6'.…”

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confidence: 99%

The auto‐oxidation of tetrahydrobiopterin

Davis

Kaufman

Milstien

1988

European Journal of Biochemistry

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The product of the aerobic oxidation of tetrahydrobiopterin, quinonoid dihydrobiopterin, is unstable and rapidly rearranges to form a 7,s-dihydropteridine. Kaufman [Kaufman, S. (1967) J. Biol. Chem. 242,3934-39431 identified the stable product produced in 0.1 M phosphate pH 6.8, as 7,s-dihydrobiopterin. However, Armarego et al. [Armarego, W. L. F., Randles, D. and Taguchi, H. (1983) Eur. J. Biochem. 135 393-4031 questioned this assignment because they found that the dihydroxypropyl group on C-6 was eliminated and 7,s-dihydropterin was the predominant product when the aerobic oxidation was performed in 0.1 M Tris pH 7.6. In the present study we demonstrate that the rearrangement of the unstable quinonoid dihydrobiopterin results in a mixture of these two 7,8-dihydropteridines at neutral pH, 25 "C. Furthermore, we find that the loss or retention of the alkyl sidechain is not solely dependent on the pH of the reaction mixture, as was previously assumed by Armarego et al., but rather is strongly influenced by the temperature and the type of buffer. In addition, we describe a new method for quantifying the relative amounts of these two 7,8-dihydropteridines in mixtures of unknown concentrations. This method relies on multicomponent analysis of second derivative spectra and results in values which agree with the concentrations determined directly by HPLC.A reduced pteridine cofactor is necessary for phenylalanine hydroxylase to catalyze the conversion of phenylalanine to tyrosine [l]. The natural cofactor for this enzyme (as well as for tyrosine hydroxylase [2-51 and tryptophan hydroxylase [6, 71 is tetrahydrobiopterin, (BH,) [S]. During the hydroxylation reaction at neutral pH, there is a stoichiometric oxidation of BH4 to quinonoid dihydrobiopterin (qBH2) [9, 101. This latter species can then be reduced back to BH4 by the NADH-dependent enzyme, dihydropteridine reductase [ l l -151. qBH2 is unstable and, in the absence of the reductase, rapidly rearranges to a 7,s-dihydropteridine [9]. It was concluded that 7,s-dihydrobiopterin (BH,) was the product of the tautomerization of qBH2 [16,17] because when the auto-oxidation of BH4 was performed in 0.1 M phosphate pH 6.8 at ambient temperature, the ultraviolet spectrum of the product was identical to that of authentic BH2 [16]. This conclusion was also strongly supported by the finding that one of the products of the aerobic oxidation of BH4 in phosphate buffer was a substrate for sepiapterin reductase, an enzyme that catalyzes the NADP-dependent oxidation of the 2'-hydroxyl group on the side-chain of BH2 [16].However, it has become apparent that the unstable qBH2 does not always isomerize to BH,. Whereas the oxidation of Correspondence to

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“…NH 2 is described as an antioxidant and as strong radical scavenger (5,8). At neutral pH neopterin remains stable, whereas NH 2 is oxidized to 7,8-dihydroxanthopterin in oxygen saturated solution (17). It has been shown that the scavenging properties of NH 2 in reactions with H 2 O 2 is based on its oxidation to 7,8-dihydroxanthopterin and xanthopterin (12).…”

Section: Discussionmentioning

confidence: 99%