The Characterization of Cytochrome P-450 by Ryo Sato
Year: 2006
Type: article
Abstract: The Carbon Monoxide-binding Pigment of Liver Microsomes. I. Evidence for Its Hemoprotein Nature (Omura, T., and Sato, R. (1964) J. Biol. Chem. 239, 2370–2378) The Carbon Monoxide-binding Pigment of Liver Microsomes. II. Solubilization, Purification, and Properties (Omura, T., and Sato, R. (1964) J. Biol. Chem. 239, 2379–2385) Ryo Sato (1923–1996) was trained in organic chemistry and biochemistry at the University of Tokyo. He did a postdoctoral fellowship at Nagoya University and became interested in the cytochromes when he discovered the role of cytochrome b1 in anaerobic nitrate respiration of Escherichia coli. In 1951, he was appointed Assistant Professor at Kanazawa University. Six years later he joined the Institute for Protein Research at the University of Osaka, where he eventually advanced to the position of Director in 1985. In 1958, shortly after Sato had joined the University of Osaka, Martin Klingenberg published a paper reporting the presence of a redox pigment in liver microsomes (1Klingenberg M. Pigments of rat liver microsomes.Arch. Biochem. Biophys. 1958; 75: 376-386Crossref PubMed Scopus (441) Google Scholar). The reduced form of the pigment bound carbon monoxide and exhibited a difference spectrum with an intense peak at 450 nm. However, because of its unprecedented spectral properties, Klingenberg was unable to explain the chemical nature of the pigment. A few months later another paper on the pigment was published (2Garfinkel D. Studies on pig liver microsomes. I. Enzymatic and pigment composition of different microsomal fractions.Arch. Biochem. Biophys. 1958; 77: 493-509Crossref PubMed Scopus (274) Google Scholar), but its nature still remained elusive. Because of his interest in cytochromes, Sato was curious about the pigment but was unable to study it because he didn't have a spectrophotometer powerful enough to measure difference spectra in turbid microsomal suspensions. Fortunately, the Institute decided to purchase a Cary 14 recording spectrophotometer for joint use, and in April 1960 Sato was able to start work on the pigment. He was aided by Tsuneo Omura, who had just joined his group. Together they were able to show that the pigment was reducible by NADH and NAD(P)H and was highly autoxidizable, but they could still not determine its chemical nature. Then, after hearing a lecture by Kozo Kaziro from which he learned that ethyl isocyanide causes a spectral change in hemoglobin, Sato decided to examine the effect of the compound on the pigment. To his surprise, he found that ethyl isocyanide induced a spectral change that was characteristic of a hemoprotein. Omura did some competition experiments and showed that ethyl isocyanide and carbon monoxide bound to the same entity in the microsomes. He was also able to show that addition of heated snake venom and deoxycholate to the microsomes converted the pigment to an alternate reduced form, which absorbs at 420 nm instead of 450 nm. Sato and Omura published a preliminary report on the pigment, which they called “P-450” (for “pigment absorbing at 450 nm”), and described it as a new cytochrome (3Omura T. Sato R. A new cytochrome in liver microsomes.J. Biol. Chem. 1962; 237: 1375-1376Abstract Full Text PDF PubMed Google Scholar). Two years later they published two papers in the Journal of Biological Chemistry (JBC) reporting the full account of their work. Both papers are reprinted here as JBC Classics. The first Classic gives a detailed description of the above-mentioned investigations on rabbit liver microsomes and crude microsomal digests, which led Sato and Omura to postulate that P-450 is a hemoprotein. Since its publication, this paper has been cited in more than 6305 publications. Sato explained this phenomenon by saying, “Why has our paper been cited so many times? The reason is quite simple. Since our paper gives an extinction coefficient increment for P-450 in the CO-difference spectrum and this value is essential for determination of P-450, most workers have to cite the paper. The increase in the number of citations to our paper simply reflects the increase in the number of publications in the field on P-450 (4Sato R. Cytochrome P-450: an inconspicuous start.Curr. Contents. 1991; 8: 9Google Scholar).” The second JBC Classic, published back-to-back with the first, deals with the solubilization and purification of the pigment. Prior to the publication of this paper, P-450 had not yet been separated from the microsomal hemoprotein cytochrome b5, and the properties of P-450 and P-420 had only been studied by means of difference spectrophotometry. Sato and Omura succeeded in separating the two microsomal hemoproteins using a technique they called “fractional solubilization.” They first treated the microsomes with steapsin, which liberated cytochrome b5 into solution but left P-450 attached to the undigested residue they called “CO-binding particles.” Treatment of these particles with heated snake venom, or deoxycholate, converted all the hemoprotein into the P-420 form and brought it into solution. The purified preparations possessed spectral properties characteristic of a hemoprotein, thus confirming the nature of the pigment suggested in the first paper. Within a few years of when these papers were published, interest in P-450 grew rapidly. Since then, the number of P-450s cloned and sequenced has expanded to over 500. The many roles of P-450s in biology and medicine now include drug metabolism, fatty acid omega oxidation, activation of chemicals initiating carcinogenesis, and vitamin D hydroxylation reactions as well as the enzymatic reactions associated with the pigments of flowers, the flavorings of spices, and the resistance to pesticides by insects. In 1985 Sato received the Imperial Award of the Japan Academy from Emperor Hirohito for his contributions to science. He served as an Editor of Archives of Biochemistry and Biophysics from 1970 to 1980 and served to foster the journal as the one of choice for publication by authors of many studies characterizing the cytochromes P-450. Tsuneo Omura, Sato's coauthor on the two Classics, is currently a professor emeritus at Kyushu University in Fukuoka. He received his B.S. degree (1953) and his D.Sc. (1962) from the University of Tokyo and was an instructor at Shizuoka University from 1953 to 1960. After working in Sato's laboratory, Omura became an assistant professor at Osaka University (1960) and remained there for 10 years. In 1970 he joined the faculty at Kyushu where he has spent the remainder of his career.
Source: Journal of Biological Chemistry
Language: English
Subfield: Inorganic Chemistry
Field: Chemistry
Domain: Physical Sciences
Open Access status: hybrid
Estimated APC paid: $3,430
OpenAlex ID: W1599381479