Fluorine-19 nuclear magnetic resonance investigation of the noncovalent and covalent binary complexes of 5-fluorodeoxyuridylate and Lactobacillus casei thymidylate synthetase

Lewis, Charles A.; Ellis, Paul D.; Dunlap, R. Bruce

doi:10.1021/bi00542a018

Cited by 40 publications

(25 citation statements)

References 25 publications

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“…The results in Table 1 for dUMP binding are consistent with these findings even when U89 is added. The binding of FdUMP clearly reveals that both active sites contribute to its binding but in an asymmetric mannerthat is, binding to one site is preferred over the other (24,37,38). However, in the presence of U89 the binding is reduced to a single site that is comparable to that obtained with dUMP, dGMP, and GMP.…”

Section: Discussionmentioning

confidence: 64%

Promotion of purine nucleotide binding to thymidylate synthase by a potent folate analogue inhibitor, 1843U89.

Weichsel

Montfort

Cieśla

et al. 1995

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

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A folate analogue, 1843U89 (U89), with potential as a chemotherapeutic agent due to its potent and specific inhibition of thymidylate synthase (TS; EC 2.1.1.45), greatly enhances not only the binding of 5-fluoro-2'-deoxyuridine 5'-monophosphate (FdUMP) and dUMP to Escherichia coli TS but also that of dGMP, GMP, dIMP, and IMP. Guanine nucleotide binding was first detected by CD analysis, which revealed a unique spectrum for the TS-dGMP-U89 ternary complex. The quantitative binding of dGMP relative to GMP, FdUMP, and dUMP was determined in the presence and absence of U89 by ultrafiltration analysis, which revealed that although the binding of GMP and dGMP could not be detected in the absence of U89 both were bound in its presence. The Kd for dGMP was about the same as that for dUMP and FdUMP, with binding of the latter two nucleotides being increased by two orders of magnitude by U89. An explanation for the binding of dGMP was provided by x-ray diffraction studies that revealed an extensive stacking interaction between the guanine of dGMP and the benzoquinazoline ring of U89 and hydrogen bonds similar to those involved in dUMP binding. In addition, binding energy was provided through a water molecule that formed hydrogen bonds to both N7 ofdGMP and the hydroxyl of Tyr-94. Accommodation of the larger dGMP molecule was accomplished through a distortion of the active site and a shift of the deoxyribose moiety to a new position. These rearrangements also enabled the binding of GMP to occur by creating a pocket for the ribose 2' hydroxyl group, overcoming the normal TS discrimination against nucleotides containing the 2' hydroxyl.

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Section: Discussionmentioning

confidence: 64%

Promotion of purine nucleotide binding to thymidylate synthase by a potent folate analogue inhibitor, 1843U89.

Weichsel

Montfort

Cieśla

et al. 1995

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

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“…The similarities between the L. casei and E. coli TS-FdUMP binary complex 19F NMR spectra showed that the E. coli enzyme, like the L. casei enzyme, bound FdUMP as an equilibrium mixture of covalent and noncovalent forms. Furthermore, the downfield narrow resonances at 0.0 and -0.7 ppm have been assigned to the S'-phosphate isomer of FdUMP and the 3'-phosphate isomer of FdUMP, both free in solution, respectively (Lewis et al, 1980) resonance intensity loss at -34.5 ppm as a result of signal digitizer dynamic range limitations. The binary complex sample was converted to pseudotemary complex by the addition ofHtfolate (Figure 2B).…”

Section: Resultsmentioning

confidence: 99%

“…The first evidence that TS could form a covalent binary complex with FdUMP was provided by I9F NMR 0006-2960/93/0432-9274$04.00/0 (Lewis et al, 1980). These results were confirmed through a nitrocellulose filter technique (Ahmed et al, 1985) and by a trichloroacetic acid precipitation procedure (Moore et al, 1984).…”

mentioning

confidence: 94%

Substitution of glutamine for glutamic acid-58 in Escherichia coli thymidylate synthase results in pronounced decreases in catalytic activity and ligand binding

Zapf¹,

Weir²,

Emerick³

et al. 1993

Biochemistry

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The recent determination of the crystal structure of Escherichia coli thymidylate synthase (TS) [Matthews et al. (1989) J. Mol. Biol. 205, 449-454] has implicated the glutamic acid residue at position 58 in a mechanistic role which could involve the interaction of its gamma-carboxyl side chain with the nucleotide substrate and/or the folate cofactor. The site-specific mutagenesis of Glu-58 to Gln-58 in E. coli TS provided the opportunity to explore its functional role in activity and binding. When profiled by the spectrophotometric and tritium release assays, the 370- and 760-fold decreases, respectively, in kcat and the elevated Km values for the Gln-58 mutant enzyme indicated a significant involvement of Glu-58 in substrate binding and turnover. The apparent dissociation constant for the covalent FdUMP-enzyme binary complex was 30 microM, which is 5-fold higher than that found for the wild-type enzyme, while the inhibitory ternary complex apparent dissociation constants for FdUMP and CH2H4folate for the Gln-58 enzyme were 10- and 60-fold higher, respectively, than those for the wild-type enzyme under saturating conditions. The extent of covalent FdUMP binding to the Gln-58 enzyme was reduced from 1.5 to 0.7 per dimer in the inhibitory ternary complex but only from 0.7 to 0.5 per dimer in the binary complex of the Gln-58 enzyme. The usual 2.1-fold enhancement of FdUMP binding to wild-type TS in the presence of CH2H4folate was not observed for the Gln-58 enzyme.(ABSTRACT TRUNCATED AT 250 WORDS)

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“…Noticeably, zinc oxide nanoparticles (ZnONPs), as one of the most important metal oxide nanoparticles, have been used as therapeutic agents in many biomedical applications due to their unique physical and chemical properties 17 . Further, the anti-inflammatory and antidiabetic effects of ZnONPs were reported previously as a novel agent to deliver zinc 18 , 19 .…”

Section: Introductionmentioning

confidence: 99%

Ameliorative effect of curcumin and zinc oxide nanoparticles on multiple mechanisms in obese rats with induced type 2 diabetes

Abdulmalek

Eldala

Awad

et al. 2021

Sci Rep

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The present study was carried out to investigate the therapeutic effect of synthesized naturally compounds, curcumin nanoparticles (CurNPs) and metal oxide, zinc oxide nanoparticles (ZnONPs) on a high-fat diet (HFD)/streptozotocin (STZ)-induced hepatic and pancreatic pathophysiology in type 2 diabetes mellitus (T2DM) via measuring AKT pathway and MAPK pathway. T2DM rats were intraperitoneally injected with a low dose of 35 mg/kg STZ after being fed by HFD for 8 weeks. Then the rats have orally received treatments for 6 weeks. HFD/STZ-induced hepatic inflammation, reflected by increased phosphorylation of p38-MAPK pathway’s molecules, was significantly decreased after nanoparticle supplementation. In addition, both nanoparticles significantly alleviated the decreased phosphorylation of AKT pathway. Further, administration of ZnONPs, CurNPs, conventional curcumin, and ZnSO4 (zinc sulfate), as well as metformin, effectively counteracted diabetes-induced oxidative stress and inflammation in the internal hepatic and pancreatic tissues. Based on the results of the current study, ZnONPs and CurNPs could be explored as a therapeutic adjuvant against complications associated with T2DM. Both nanoparticles could effectively delay the progression of several complications by activating AKT pathway and down-regulating MAPK pathway. Our findings may provide an experimental basis for the application of nanoparticles in the treatment of T2DM with low toxicity.

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Fluorine-19 nuclear magnetic resonance investigation of the noncovalent and covalent binary complexes of 5-fluorodeoxyuridylate and Lactobacillus casei thymidylate synthetase

Cited by 40 publications

References 25 publications

Promotion of purine nucleotide binding to thymidylate synthase by a potent folate analogue inhibitor, 1843U89.

Promotion of purine nucleotide binding to thymidylate synthase by a potent folate analogue inhibitor, 1843U89.

Substitution of glutamine for glutamic acid-58 in Escherichia coli thymidylate synthase results in pronounced decreases in catalytic activity and ligand binding

Ameliorative effect of curcumin and zinc oxide nanoparticles on multiple mechanisms in obese rats with induced type 2 diabetes

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