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Human oxyhemoglobin showed a biphasic autoxidation curve containing two rate constants, i.e. k f for the fast autoxidation due to the ␣ chains, and k s for the slow autoxidation of the ␤ chains, respectively. Consequently, the autoxidation of the HbO 2 tetramer produces two different curves from the pH dependence of k f and k s . The analysis of these curves revealed that the ␤ chain of the HbO 2 tetramer does not exhibit any protoncatalyzed autoxidation, unlike the ␣ chain, where a proton-catalyzed process involving the distal histidine residue can play a dominant role in the autoxidation rate. When the ␣ and ␤ chains were separated from the HbO 2 tetramer, however, each chain was oxidized much more rapidly than in the tetrameric parent. Moreover, the separated ␤ chain was recovered completely to strong acid catalysis in its autoxidation rate. These new findings lead us to conclude that the formation of the ␣ 1 ␤ 1 contact produces in the ␤ chain a conformational constraint whereby the distal histidine at position 63 is tilted away slightly from the bound dioxygen, preventing the proton-catalyzed displacement of O 2. by a solvent water molecule. The ␤ chains have thus acquired a delayed autoxidation in the HbO 2 tetramer.The reversible and stable binding of molecular oxygen to the heme iron(II) is the basis of hemoglobin function. Consequently, much attention has been directed to the elucidation of the molecular mechanism of cooperative oxygen binding to the hemoglobin tetramer (1). However, the oxygenated form of hemoglobin, as well as of myoglobin, is known to be oxidized easily to the ferric(III) met form, which cannot bind molecular oxygen and is therefore physiologically inactive, with generation of the superoxide anion (2-5).To this autoxidation reaction, it has been widely accepted that hemoglobin is much more resistant as compared with myoglobin. Unlike myoglobin, Mansouri and Winterhalter (6) reported that oxyhemoglobin (HbO 2 ) showed a biphasic autoxidation reaction with a fast and a slow component. They also demonstrated that the ␣ chain was oxidized more rapidly than the ␤ chain in hemoglobin tetramer. At the same time, however, there have been a number of reports that such a rate difference was not observed between the ␣ and ␤ chains in the autoxidation reaction of HbA (7-9). Rather, Zhang et al. (9) showed that the rate of autoxidation was markedly enhanced when the HbO 2 tetramer dissociates into ␣␤ dimers. To clarify these discrepancies, we have recently examined systematically the effect of hemoglobin concentration on the autoxidation rate at several different values of pH, and found that human HbO 2 exhibits a biphasic autoxidation curve only in the pH range from neutral to acidic (5). By dissociation of tetramers into ␣␤ dimers, the rate of autoxidation for the fast component (due to the ␣ chain) was also found to increase markedly at the acidic pH, but the addition of 2,3-diphosphoglyceric acid offered no significant effect on the increment of the autoxidation rate (5).In the present pape...

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Nuclear receptor family proteins are structurally related transcription factors activated by specific lipophilic compounds. Because they are activated by a variety of hormonal molecules, including retinoic acid, vitamin D, and steroid hormones, they are assumed to be promising targets for clinical drugs. We previously found that one ascochlorin (1) derivative, 4-O-carboxymethyl-ascochlorin (2), is a potent agonist of peroxisome proliferator activated receptor gamma (PPARgamma). Here, we synthesized derivatives of 1, designated as a lead compound, to create new modulators of nuclear hormone receptors. Two derivatives, 4-O-carboxymethyl-2-O-methylascochlorin (9) and 4-O-isonicotinoyl-2-O-methylascochlorin (10), showed improved agonistic activity for PPARgamma and induced differentiation of a progenitor cell line, C3H10T1/2. We also found that 1, dehydroascofuranon (29), and a 2,4-O-diacetyl-1-carboxylic acid derivative of 1 (5) specifically activated estrogen receptors, PPARalpha, and an androgen receptor. All of the derivatives (1-29) activated the pregnane X receptor. These results suggest that the chemical structure of 1 is useful in designing novel modulators of nuclear receptors.