Bongkrekic acid

Bongkrekic acid (BKA), isolated from the bacterium Burkholderia cocovenenans, is an inhibitor of adenine nucleotide translocator, which inhibits apoptosis, and is thus an important tool for the mechanistic investigation of apoptosis. An efficient total synthesis of BKA has been achieved by employing a three-component convergent strategy based on Kocienski-Julia olefination and Suzuki-Miyaura coupling. It is noteworthy that segment B has been prepared as a new doubly functionalized coupling partner, which contributes to shortening of the number of steps. Torquoselective olefination with an ynolate has also been applied for the efficient construction of an unsaturated ester. Furthermore, it is revealed that 1-methyl-2-azaadamantane N-oxyl is an excellent reagent for final oxidation to afford BKA in high yield. Based on the total synthesis, several BKA analogues were prepared for structure-activity relationship studies, which indicated that the carboxylic acid moieties were essential for the apoptosis inhibitory activity of BKA.

Bongkrekic acid, isolated from Burkholderia cocovenenans, is known to specifically inhibit the mitochondrial ADP/ATP carrier. However, the manner of its interaction with the carrier remains elusive. In this study, we tested the inhibitory effects of 17 bongkrekic acid analogues, derived from the intermediates obtained during its total synthesis, on the mitochondrial ATP/ATP carrier. Rough screening of these chemicals, performed by measuring their inhibitory effects on the mitochondrial ATP synthesis, revealed that 4 of them, KH-1, KH-7, KH-16, and KH-17, had moderate inhibitory effects. Further characterization of the actions of these 4 analogues on mitochondrial function showed that KH-16 had moderate; KH-1 and KH-17, weak; and KH-7, negligible side effects of both permeabilization of the mitochondrial inner membrane and inhibition of the electron transport, indicating that only KH-7 had a specific inhibitory effect on the mitochondrial ADP/ATP carrier.

Giant seaperch iridovirus (GSIV) induces cell death by an unknown mechanism. We postulated that this mechanism involves mitochondria-mediated cell death. Cell viability assays revealed a steady increase in dead grouper fin cells (GF-1) after GSIV infection, from 11% at 2 days post-infection (dpi) to 67% at 5dpi. Annexin V/PI staining revealed GSIV infection induced apoptosis in a steadily increasing fraction of cells, from 4% at 1dpi to 29% at 5dpi. Furthermore, post-apoptotic necrosis was apparent at 4 and 5dpi in the late replication stage. In the early replication stage, JC-1 dye revealed mitochondrial membrane potential (ΔΨm) loss in 42% of infected cells at 1dpi, increasing to 98% at 3dpi. Phosphatidylserine (PS) exposure and loss of ΔΨm from apoptosis/necrosis was attenuated by treatment with the adenine nucleotide translocase inhibitor bongkrekic acid (BKA) and the protein synthesis inhibitor cyclohexamide (CHX). These data suggest GSIV induces GF-1 apoptotic/necrotic cell death through pathways that require newly synthesized protein and involve the mitochondrial function.