Peroxidase activity of myoglobin is enhanced by chemical mutation of heme-propionates

Takashi Hayashi, Yutaka Hitomi, Tsutomu Ando, Tadashi Mizutani, Yoshio Hisaeda, Susumu Kitagawa, Hisanobu Ogoshi

Research output: Contribution to journalArticle

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Abstract

Peroxidase activity of a myoglobin reconstituted with a chemically modified heme 1 is reported. The heme 1 bearing a total of eight carboxylates bound to the terminal of propionate side chains is incorporated into apomyoglobin from horse heart to obtain a new reconstituted myoglobin, rMb(1), with a unique binding domain structure. The UV-vis, CD, and NMR spectra of rMb(1) are comparable with those of native myoglobin, nMb. The mixing of rMb(1) with hydrogen peroxide yields a peroxidase compound II-like species, rMb(1)-II, since the spectrum of rMb(1)-II is identical with that observed for nMb. Stoichiometric oxidation of several small molecules by rMb(1)-II, demonstrates the significant reactivity. (i) The oxidation of cationic substrate such as [Ru(NH3)6]2+ by rMb(1)-II is faster than that observed for oxoferryl species of nMb, nMb-II. (ii) Anionic substrates such as ferrocyanide are unsuitable for the oxidation by rMb(1)-II. (iii) Oxidations of catechol, hydroquinone, and guaiacol are dramatically enhanced by rMb(1)-II (14-32-fold) compared to those observed for nMb-II. Thus, the chemical modification of heme-propionates can alter substrate specificity. Steady-state kinetic measurements indicate that both the reactivity and substrate affinity toward guaiacol oxidation by rMb(1) are improved, so that the specificity, k(cat)/K(m), is 13-fold higher than that in nMb. This result strongly suggests that the artificially modified heme-propionates may increase the accessibility of neutral aromatic substrates to the heme active site. The present work demonstrates that the chemical mutation of prosthetic group is a new strategy to create proteins with engineered function.

Original languageEnglish
Pages (from-to)7747-7750
Number of pages4
JournalJournal of the American Chemical Society
Volume121
Issue number34
DOIs
Publication statusPublished - Sep 1 1999

Fingerprint

Myoglobin
Propionates
Heme
Peroxidase
Guaiacol
Oxidation
Mutation
Substrates
Substrate Specificity
Bearings (structural)
Hydrogen Peroxide
Horses
Catalytic Domain
Chemical modification
Prosthetics
Hydrogen peroxide
Nuclear magnetic resonance
Proteins
Molecules
Kinetics

All Science Journal Classification (ASJC) codes

  • Catalysis
  • Chemistry(all)
  • Biochemistry
  • Colloid and Surface Chemistry

Cite this

Peroxidase activity of myoglobin is enhanced by chemical mutation of heme-propionates. / Hayashi, Takashi; Hitomi, Yutaka; Ando, Tsutomu; Mizutani, Tadashi; Hisaeda, Yoshio; Kitagawa, Susumu; Ogoshi, Hisanobu.

In: Journal of the American Chemical Society, Vol. 121, No. 34, 01.09.1999, p. 7747-7750.

Research output: Contribution to journalArticle

Hayashi, T, Hitomi, Y, Ando, T, Mizutani, T, Hisaeda, Y, Kitagawa, S & Ogoshi, H 1999, 'Peroxidase activity of myoglobin is enhanced by chemical mutation of heme-propionates', Journal of the American Chemical Society, vol. 121, no. 34, pp. 7747-7750. https://doi.org/10.1021/ja9841005
Hayashi, Takashi ; Hitomi, Yutaka ; Ando, Tsutomu ; Mizutani, Tadashi ; Hisaeda, Yoshio ; Kitagawa, Susumu ; Ogoshi, Hisanobu. / Peroxidase activity of myoglobin is enhanced by chemical mutation of heme-propionates. In: Journal of the American Chemical Society. 1999 ; Vol. 121, No. 34. pp. 7747-7750.
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