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

86 Citations (Scopus)

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

All Science Journal Classification (ASJC) codes

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

Fingerprint Dive into the research topics of 'Peroxidase activity of myoglobin is enhanced by chemical mutation of heme-propionates'. Together they form a unique fingerprint.

  • Cite this