Design and synthesis of semi-artificial myoglobin possessing DNA-binding peptides on heme propionates

Seiji Sakamoto, Kazuaki Kudo

Research output: Contribution to journalArticlepeer-review

14 Citations (Scopus)

Abstract

Peptides possessing the DNA-binding capability were successfully introduced into myoglobin by a cofactor-reconstitution strategy. We have designed and synthesized an artificial iron-protoporphyrin IX (heme)-peptide conjugate, Heme(br) 2, which has covalently conjugated GCN4 basic region peptides at the heme propionate groups as an artificial DNA-binding site. The peptide Heme(br) 2 was inserted into apomyoglobin to give a peptide-conjugated myoglobin, Mb(br) 2. The UV-vis and circular dichroism spectra of Mb(br) 2 were comparable to those of native myoglobin, suggesting that the heme environment and three-dimensional structure of Mb(br) 2 were almost identical to those of native myoglobin. The size exclusion chromatography indicated that the Mb(br) 2 existed as a monomeric form in an aqueous solution and interacted with its target DNA sequence [c-AMP responsive element (CRE)]. The fluorescence study also revealed that the Mb(br) 2 bound CRE DNA effectively with 1/1 stoichiometry [affinity constant; 8.4 (±0.8) × 10 7 M -1]. Additionally, the CRE DNA-binding enhanced the peroxidase-like activity of Mb(br) 2, possibly due to the partial structural perturbation around the heme active-site. These results demonstrate that the chemical modification of prosthetic group with functional peptides can provide a new strategy for the design of semi-artificial proteins with engineered function.

Original languageEnglish
Pages (from-to)1749-1756
Number of pages8
JournalBulletin of the Chemical Society of Japan
Volume78
Issue number10
DOIs
Publication statusPublished - 2005 Dec 1

ASJC Scopus subject areas

  • Chemistry(all)

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