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. 2007 Sep 19;129(37):11334-5.
doi: 10.1021/ja074446s. Epub 2007 Aug 23.

Direct spectroscopic evidence for a high-spin Fe(IV) intermediate in tyrosine hydroxylase

Bekir E Eser  1 , Eric W Barr, Patrick A Frantom, Lana Saleh, J Martin Bollinger Jr, Carsten Krebs, Paul F Fitzpatrick
Affiliations

Direct spectroscopic evidence for a high-spin Fe(IV) intermediate in tyrosine hydroxylase

Bekir E Eser et al. J Am Chem Soc. .
No abstract available

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Figures

Figure 1
Figure 1
4.2-K Mössbauer spectra of the reaction at 5 °C of the TyrH•Fe(II) •6-MePH4•Tyr complex (2.15 mM TyrH, 1.95 mM Fe(II), 3.7 mM 6-MePH4 and 3.7 mM Tyr in 200 mM Hepes, 10% glycerol, 0.1 M KCl at pH 7.5) with 1.9 mM oxygen-containing buffer in a ratio of 1:2. Reaction times and magnetic field strengths are as indicated. Left panel: Spectra (hashed marks) at various reaction times. The solid lines are quadrupole doublet simulations of the Fe(IV) intermediate (δ = 0.25 mm/s and ΔEQ = 1.27 mm/s). Right panel: Deconvolution of the spectrum of the 20-ms sample in zero-field (top panel) and an 8-T field (bottom panel). The spectrum of the anaerobic control scaled to 60% of the total intensity is shown as a solid line overlaid with the raw data. The resulting difference spectra (hashed marks) can be simulated with spin Hamiltonian simulations of the Fe(IV) intermediate (24% intensity) with the following parameters: S = 2, D = 12.5 cm−1, E/D = 0.05, δ = 0.25 mm/s, ΔEQ = −1.27 mm/s, η = −0.5, A/gNβN = (−18.0, −18.0, −31.0) T.
Figure 2
Figure 2
Comparison of time courses for Fe(IV)O formation and decay (diamonds) and for DOPA formation (circles). DOPA was quantified by rapid-quench of the reaction at 5 °C of the complex of 500 μM TyrH, 480 μM Fe(II), 1 mM Tyr and 2 mM 6-MePH4 with an equal volume of 1.9 mM oxygen-containing buffer. The lines are simulations using the mechanism of Scheme 2 and values of k1 and k2 of 24 and 35 s−1, respectively, assuming that 80% of the enzyme complex is active.
Scheme 1
Scheme 1
Scheme 2
Scheme 2
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References

    1. Abbreviations used: TyrH, tyrosine hydroxylase; Tyr, tyrosine; DOPA, dihydroxyphenylalanine; 6-MePH4, 6-methyl tetrahydropterin; 4a-HOPH3, 4a-hydroxypterin.

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