以前の日本語版研究内容はこちら。

Hemoprotein Engineering

Hemoproteins, which have iron porphpyrins as a prosthetic group, show a variety of functions derived from the unique combination of the heme and protein matrix. Thus, it is of considerable interest to engineer the function of the hemoproteins. To modify the hemoproteins, at least there are three methods as follows. Our group mainly focuses on the third method, "Insertion of an artificially created heme into an apoprotein" to yield a reconstituted protein to understand the function and/or enhance the reactivity.1 1) Acc. Chem. Res., 35, 35-43 (2002).

Enhancement of Hemoprotein Reactivity

In a series of hemoproteins, heme b (protoporphyrin IX iron complex), is bound in the heme pocket via multiple non-covalent interactions. To enhance and/or modify the hemoprotein functions, we have prepared several functionalized artificial heme derivatives or analogs (isomers) and replaced the native heme with them as shown in the following scheme. The obtained reconstituted hemoproteins exhibit unique physicochemical or enzymatic properties: extremely O2 affinity,2-5 remarkable peroxidase activity,6-9 reductive O2 activation,10 and interprotein electron transfer. Target proteins for our research are myoglobin, hemoglobin, horseradish peroxidase, cytochrome b562, cytochrome c, cytochrome P450cam, heme oxygenase etc. and those mutants. 2) J. Am. Chem. Soc., 124, 11226-11227 (2002). 3) J. Am. Chem. Soc.,126, 16007-16017 (2004). 4) J. Am. Chem. Soc.,127, 56-57 (2005). 5) Inorg. Chem.,44, 9391-9396 (2005). 6) J. Am. Chem. Soc.,121, 7747-7750 (1999). 7) J. Am. Chem. Soc.,126, 436-437 (2004). 8) Inorg. Chem., 45, 10530-10536 9) J. Am. Chem. Soc.,129, 12906-12907 (2007). 10) J. Am. Chem. Soc.,124, 11234-11235 (2002).

Molecular Recognition on the Protein Surface

Biological communication on the protein or cell surface is one of the attractive phenomena. Thus, it is of particular interest to understand the physicochemical properties of protein-protein interaction. Our group has first proposed a modified myoglobin which has an anionic patch on the protein surface to make a complex with a positively charged cytochrome c.11-12 The negatively or positively charged protein is prepared by inserting the heme with modified propionate side chains having multiple carboxylate or ammonium groups, respectively, into the apohemoprotein. The complex between negatively charged zinc myoglobin and cytchrome c is a good model for an interprotein electron transfer in mitochondrial respiratory system.13 In addition, zinc myoglobin with the anionic patch on the protein surface can act as a photocatalyst which can bind positively charged methyl viologen, because the obtained reduced viologen is a good electron donor to generate H2 gas in the presence of Pt colloid.14 Furthermore, we obtained a reconstituted myoglobin with sugar moieties at the terminal of two heme-propionates. The glycomyoglobin gives a unique complex with a sugar binding protein such as penut lectin via the terminal carbohydrate units as an interface.15 11) Chem. Soc. Rev.,126, 355-364 (1997). 12) J. Am. Chem. Soc.,120, 4910-4915 (1998). 13) Angew. Chem., Int. Ed.,113, 1132-1135 (2001). 14) Chem. Commun., 31, 3684-3686, (2008). 15) Chem. Commun., 29, 3131-3133 (2006).

Supramolecular Protein Polymer

Modifying the architecture of a supramolecular polymer is a promising tactic to mimic a biological system and construct bionanomaterials. We found that the heme-heme pocket interaction with an affinity of 1010 - 1015 M-1 serves as a new way to prepare an attractive supramolecular hemoprotein polymer as follows. At the first stage, we introduced iron porphyrin derivatives onto the thiol group of the cytochrome b562 mutant (H63C). The removal of the native heme from the protein induces the attached external heme to bind into the heme pocket intermolecularly. The subsequent heme-heme pocket interaction produces a unique submicrosized "1D hemoprotein fiber" or "hemoprotein donut" containing more than 200 proteins.16-17 Furthermore, the addition of a heme triad into the hemoprotein fiber gives a beautiful "2D hemoprotein monolayer network" with the height of ca. 5 nm on the HOPG substrate. The present strategy can easily be extended to prepare many kinds of hemoprotein assemblies with various protein cluster geometries.18 16) J. Am. Chem. Soc., 129, 10326-10327 (2007). 17) Biopolymers, 3, 194-200, (2009). Cover picture 18) Angew. Chem., Int. Ed., 48, 1271-1274 (2009). Issue picture

Functional Role of Heme-Propionate Side Chain in Hemoprotein

Heme b has two propionate side chains. It has been believed that the propionate side chains play a role on the anchor of the prosthetic group heme to bind with hemoprotein interior. We propose the functional significance of those propionate side chains by reconstituted myoglobins with two one-legged hemes where one of the propionates is replaced with methyl group. In sperm whale myoglobin, it is found that the heme-6-propionate side chain stabilizes the bound dioxygen via unique hydrogen bonging network in the distal pocket.14 In monooxygenase cytochrome P450cam, the heme-6-propionate is essential for the maintenance of the enzyme activity, because the removal of the side chain converts the active enzyme into an inactive P420 species, although the crystal structure of the reconstituted protein is almost same as that of the wild type protein.20 The removal of the heme-7-propionate dramatically decreases the substrate d-camphor affinity and decelerates the catalytic hydroxylation of d-camphor.21 19) Biochemistry, 46, 9406-9416 (2007). 20) J. Am. Chem. Soc., 130, 432-433 (2008). 21) J. Am. Chem. Soc., 4, 1398-1400 (2009).

New Porphyrinoid Chemistry

Tetrapyrrolic metalloporphyrin derivatives have been widely studied as biomimetic models for hemoproteins. They are also well known as attractive catalysts in organic and coordination chemistry. We have focused on unique porphyrin isomers, particularly those such as porphycene with reduced symmetry, because the LUMO of the 18pi-macrocyclic porphycene is dramatically stabilized compared to the corresponding porphyrin. We prepared metalloporphycene having strongly electron-withdrawing trifluoromethyl groups at the pyrrole beta-positions to obtain extremely electron-deficient tetrapyrrole macrocycles.22 The unusual characteristics of porphycene and metalloporphycene species provide a stable low-valent metal complex,23,24 an interelement complex containing an M-M bond and a 20pi-nonaromatic porphycene.25 22) Org. Lett., 5, 2845-2848 (2003). 23) Inorg. Chem., 42, 7345-7347 (2003). 23) Bull. Chem. Soc. Jpn., 81, 76-83 (2008). BCSJ award 24) Org. Lett., 9, 5303-5306 (2007).