Photoinduced Singlet Electron Transfer in a Complex Formed from Zinc Myoglobin and Methyl Viologen: Artificial Recognition by a Chemically Modified Porphyrin

Abstract

One major goal of molecular recognition processes in biological systems is to understand how the chemical reaction is regulated by protein binding sites. For example, it is well known that the specific interaction of the cytochrome c-cytochrome c peroxidase complex plays an important role in longrange electron transfer (ET).' However, the noncovalent model systems in which fast ET occurs from donor to acceptor via intermolecular interaction remain limited.2 Here, we wish to report a nonmutagenic approach to the construction of the noncovalent ET model that is based on the complex of methyl viologen (2) and a myoglobin reconstituted with a modified zinc porphyrin (l*Zn) having an assembly of eight carboxylic acid groups as a specific interface. The present supramolecular system, designed to mimic the protein-protein complex of a reductoxidase, enables us to observe the fast singlet ET from zinc myoglobin to methyl viologen as a first example.Our strategy for the construction of rMb(l*Zn)-2 pairing is shown in Scheme 1.' Eight carboxylates attached to 6-and 7-propionates of mesoporphyrin IX were designed as an artificial recognition site for the cationic a~ceptor.~ Free base 1 was prepared by the coupling of mesoporphyrin and 5-aminoisophthalate condensed with two aspartic acids. The synthetic zinc porphyrin l*Zn was easily inserted into horse heart apomyoglobin by a routine m e t h~d .~-~ The characteristic visible absorption spectrum of the reconstituted protein was similar to that of reference protein reconstituted with mesoporphyrin zinc complex 3.231.Addition of 2 to the solution of rMb(l*Zn) brought about no changes in the visible absorption spectrum. In contrast, fluorescence quenching in rMb(l*Zn) clearly was observed upon addition of 2 in 10 mM phosphate buffer, pH 7.0, while reference reconstituted myoglobin, rMb(3*Zn), showed no (1) (a) Takano, T.; Dickerson, R. E. Throughout this paper, myoglobins reconstituted with l.Zn and 3*Zn are abbreviated as rMb( l-Zn) and rMb(3*Zn), respectively.(4) 5-[(tert-Butoxycarbonyl)amino]isophthaloyldiaspa~ic acid as a precursor interacts with 2 in pH 7.0 phosphate buffer solution. The affinity constant was determined to be (3.6 f 0.4) x IO2 M-' by 'H NMR measurement.(5) See supporting information for preparation details.(a) Teale, F. W. Bioclzim. Biophys. Acfu 1959,35,543. (b) Yonetani, T.; Asakura, T. J. B i d . Clzem.'1969, 244, 4580. (7) For examples of reconstituted semisynthetic myoglobin, cf.: (a) Suzuki, A.; Okuda, K.; Kawagoe, K.; Toi, H.; Aoyama, Y.; Ogoshi, H. Takimura, T.; Ohara, T.; Hitomi, Y.; Ogoshi, H. J. Chenz. Soc., Chem. Commun., in press. (e) Hayashi, T.; Hitomi, Y.; Suzuki, A.; Takimura, T.; Ogoshi, H. Chem. Lett. 1995, 91 1. 0002-7863/95/1517-11606$09.00/0 z I I,ll . a, C v) 2.5 i i .-> a, 1.5 L 0 3 c

New Functionalization of Myoglobin by Chemical Modification of Heme-Propionates

“…9b, 24 The shorter-and longer-lived components of the biphasic decay curve are assignable to the fluorescence lifetimes of the rMb(1‚Zn)-MV 2+ complex and free rMb(1‚Zn), respectively. From these data, a rate constant of the forward singlet ET was estimated to be k et ) 2.1 × 10 9 s -1 .…”

Heme-peptide complexes as peroxidase models

“…With the aim of creating such an artificial binding domain, Hayashi reconstituted Mb with two different types of chemically modified cofactors: an anionic heme with four or eight carboxylates at the terminal of the two propionate side chains [23,24] and a cationic heme with four amino groups at the end of the two propionate chains [25]. These modified Mbs form stable complexes both by electrostatic interactions with substrates carrying an opposite charge and by hydrophobic contacts [25].…”

Enhancement of enzymatic activity for myoglobins by modification of heme-propionate side chains

“…On the basis of this idea, we prepared a modified hemin 2 where a benzene moiety was introduced into each heme-propionate to form an artificial hydrophobic substrate-binding site as shown in Fig. 2 [21][22][23]. The reconstituted myoglobin with hemin 2 was characterized by UV-vis and ESImass spectroscopic methods.…”