Enzymes in the mitochondrial respiratory chain are involved in various physiological events in addition to their essential role in the production of ATP by oxidative phosphorylation. The use of specific and potent inhibitors of complex I (NADH-ubiquinone reductase) and complex III (ubiquinol-cytochrome c reductase), such as rotenone and antimycin, respectively, has allowed determination of the role of these enzymes in physiological processes. However, unlike complexes I, III, and IV (cytochrome c oxidase), there are few potent and specific inhibitors of complex II (succinate-ubiquinone reductase) that have been described. In this article, we report that atpenins potently and specifically inhibit the succinate-ubiquinone reductase activity of mitochondrial complex II. Therefore, atpenins may be useful tools for clarifying the biochemical and structural properties of complex II, as well as for determining its physiological roles in mammalian tissues. T he use of specific and potent inhibitors of respiration has enabled the investigation of how the respiratory enzymes function in physiological processes. However, unlike other enzyme complexes in the respiratory chain, there has been a lack of potent and specific inhibitors of complex II [succinateubiquinone reductase (SQR)]. Although carboxin (5,6-dihydro-2-methyl-N-phenyl-1,4-oxathiin-3-carboxamide), TTFA [4,4,4-trifluoro-1-(2-thienyl)-1,3-butanedione], and HQNO (2-heptyl-4-hydroxyquinoline N-oxide) have long been known as complex II inhibitors and have been used extensively to elucidate the structure-function relationships of complex II, rather higher concentration is required for the inhibition (1). This result has hampered the study of the structure-function relationship of the complex II enzyme, as well as its roles in physiological processes.Complex II catalyzes the oxidation of succinate in the inner membrane of mitochondria and in the cytoplasmic membrane of bacteria (1-3). In addition to its function as a dehydrogenase in the respiratory system, complex II plays an important role in the tricarboxylic acid cycle. Mitochondrial complex II is an integral membrane protein consisting of four subunits (Fig. 1). The largest subunit is the 70-kDa, FAD-containing flavoprotein subunit (Fp). The dehydrogenase catalytic portion of complex II is formed by Fp and an Ϸ30-kDa iron-sulfur protein subunit (Ip) containing three different types of iron-sulfur clusters. The small hydrophobic subunits, SDHC or CybL (Ϸ15 kDa) and SDHD or CybS (Ϸ13 kDa), anchor the catalytic portion to the membrane and are also required for electron transfer to quinones. In contrast to mitochondrial complex IIs, some bacterial complex IIs contain only one larger hydrophobic polypeptide as a membrane anchor (see ref. 4 for reviews).In addition to its essential role in energy production, various recent findings suggest that mutant variants of complex II are involved in causing diverse physiological disorders. For instance, a mutation in the CybL subunit in Caenorhabditis elegans (mev-1 mutant) resu...

A new chitinase inhibitor, designated as argadin (1), was isolated from the cultured broth of a fungal strain FO-7314. The strain was identified as Clonostachys sp. from the morphological characteristics. Argadin was purified from the cultured mycelium by a combination of cation exchange, adsorption and gel filtration chromatographic methods. The structure of argadin was elucidated as cyclo(Nomega-acetyl-L-arginyl-D-prolyl-homoseryl-histidyl-L- 2-aminoadipyl) in which homoseryl gamma-methylene bonded to histidyl alpha-amino residue. The IC50 value of argadin against Lucilia cuprina (blowfly) chitinase was 150 nM at 37 degrees C and 3.4 nM at 20 degrees C. Argadin arrested the moult of cockroach larvae upon injection into the ventral abdominal part.

Novel brominated and halogen-less azaphilone (oxoisochromane) derivatives, 5-bromoochrephilone and dechloroisochromophilone IV, and known derivatives, dechloroisochromophilone III and isorotiorin, were isolated from the culture broth of a producing organism of isochromophilones I and II (azaphilones inhibiting gp120-CD4 binding), Penicillium multicolor FO-2338, fermented in a medium containing potassium bromide. Nineteen azaphilone-related compounds isolated from the above strain and from other fungi were tested for the inhibition of gp120-CD4 binding and the structure-activity relationship is discussed. Consequently, 5-bromoochrephilone is the strongest inhibitor chain of 6-oxoisochromane ring are necessary for gp120-CD4 binding.Fungal metabolites having an oxoisochromane ring system, which are called "azaphilones" after high affinity to ammonia1), have been found to possess various bioactivities. In the course of screening of microbial metabolites for the inhibition of gp120-CD4 binding, we discovered the inhibitors isochromophilones I (1) and II (2) having a 6-oxoisochromane ring, which were isolated from a culture broth of Penicillium multicolor FOphilone (3)5), sclerotiorin (4)6) and rubrorotiorin (5)6).

Newazaphilones named isochromophilones III~VI were isolated from the culture broth of Penicillium multicolor FO-3216 as inhibitors of acyl-CoA : cholesterol acyltransferase (ACAT). Their structures were elucidated by NMR and other spectroscopic analyses. The IC50 values of isochromophilones III, IV, V and VI for ACATactivity in an enzyme assay using rat liver microsomes were calculated to be 110, 50, 50 and 120fiM, respectively.

An enzyme assay for bacterial undecaprenyl pyrophosphate (UPP) synthase was performed to screen microbial culture broths for inhibitors of UPP synthase. During the course of this screening program, an EtOH extract of a rice culture of Penicillium brasilianum FKI-3368 was found to inhibit UPP synthase activity. From activity-guided purification, a new compound-designated spirohexaline was isolated together with the structurally related and known viridicatumtoxin by ethyl acetate extraction silica gel and octadecylsilane column chromatographies and high-performance liquid chromatography. The structure of spirohexaline was elucidated by spectroscopic analysis, including NMR. Spirohexaline and viridicatumtoxin have a common hexacycline structure produced by fusion of a tetracycline-type ring with a spiro-type ring. They inhibited UPP synthase activity with IC₅₀ values of 9.0 and 4.0 μM, respectively.