Effect of side‐chain structure on inhibition of yeast fatty‐acid synthase by cerulenin analogues

Abstract

Yeast fatty-acid synthase (FAS) inhibition by cerulenin analogs with varying side-chain lengths was compared with that of cerulenin, tetrahydrocerulenin and iodoacetamide. Although inhibition by cerulenin was the highest, the analogs having (E,E)-A'*" double bonds showed high inhibition. This strongly suggests that the (E, E)-d7? l o double bonds play an important role in the interaction of the inhibitors with the enzyme. It was suggested that the size of the hydrophobic cavity in the condensing enzyme terminates fatty-acid chain elongation by decreasing inhibition by the C1 analog. Like cerulenin itself, the shortest analog (C,) did not induce malonyl-CoA decarboxylase activity.Cerulenin 1 [I, 21 is an antibiotic produced by Cephalosporium caerulens [3]. It has a hydrophilic head part (C1 -C4) with a cis-epoxide moiety and a hydrophobic side chain with an (E,E)-lP-diene system (C5-Cl2) [4, 51 (Fig. 1).Cerulenin inhibits fatty-acid synthase (FAS) [6] of both multifunctional and unassociated enzymes [7 -111, except the synthase from cerulenin-producing fungi [12-141. Vance et al.[15] first reported, using the enzyme preparation from Mycobacterium phlei, that 3-oxoacyl synthase (condensing enzyme) was hghly sensitive to cerulenin. Subsequently, d'Agnelo et al. [16] confirmed, with the purified enzyme from Escherichia coli, that cerulenin binds to the cysteine at the active site of the condensing enzyme (active cysteine thiol group) and inhibits the condensation of fatty-acylated acylcarrier protein (ACP) and malonyl-ACP. They observed that inhibition of the condensing enzyme by cerulenin was irreversible, and that the molar ratio of cerulenin/enzyme was approximately 1 : 1 when inhibition approached 100%.Inhibition of FAS by cerulenin is more potent and specific [7] than that by the thiol inhibitor iodoacetamide (4), which binds to the same cysteine residue [17-191. This was demonstrated by comparison of the second-order rate constants (k,) of the reactions. The values of the rate constant k, were 88 M-' . S-' [7] and 1.0 M-' . S- ' [19] for the reaction between cerulenin and yeast FAS, and between iodoacetamide and FAS, respectively. These data indicate that yeast FAS reacts with cerulenin about 90 times faster than with iodoacetamide [7].We have already demonstrated that cerulenin reacts with the cysteine thiol group at C2 to give the adduct 2b (Fig. 1) in a model reaction between cerulenin and cysteine methyl

Structure of the Complex between the Antibiotic Cerulenin and Its Target, β-Ketoacyl-Acyl Carrier Protein Synthase

“…In comparison, binding of tetrahydrocerulenin would cost entropy, and as expected it shows more than 2 orders of magnitude less inhibitory activity (27). The influence of the length of the hydrocarbon chain, maintaining the double bond positions, has been studied using fatty acid synthase from Saccharomyces cerevisiae (28). Cerulenin (12 carbons) had the highest inhibitory activity, with slightly decreasing binding strength upon increase in chain length.…”

Inhibition of the fungal fatty acid synthase type I multienzyme complex

“…However, the 1,4-nonadienoic CER was shown to be the most effective inhibitor of S. cerevisiae FAS of 11 saturated and unsaturated CER analogs (35). In the FAS-CER complex CER C5-C8 stretches into the long KS acyl pocket, interacting with the side chains of F1279, K1585, and F1343 ( Fig.…”

“…The FabF I108 has been proposed to be responsible for the low CER sensitivity compared to FabB, which has a glycine residue at this position (28,34). Because the IC 50 value of FAS CER inhibition (35) is in the range of FabB, the FAS KS has to be able to compensate for the rearrangement of the methionine, for example, by stacking interactions to the CER C7-C8 double bond and a better fit of the rigid CER tail to the end of the acyl-binding pocket.…”

“…The influence of methionine M1251 on KS acyl binding can be deduced from a systematic study on CER inhibition of S. cerevisiae FAS with respect to the length of the CER hydrophobic tail (35). Morisaki et al found that inhibition efficiencies of CER analogs truncated at C8, C9, and C10 decrease constantly (IC 50 of 60, 170, and 300 M), whereas it is significantly increased for C11-C16 analogs (IC 50 between 30 and 4.5 M), indicating the rotational barrier of M1251 and its compensation at longer chain lengths.…”

Inhibition of InhA Activity, but Not KasA Activity, Induces Formation of a KasA-containing Complex in Mycobacteria

“…CER is a drug that is produced naturally by Cephalosporium caerulens and exhibits potent activity against a wide variety of yeast, fungi, and bacteria (53) through inhibition of both FAS-I and FAS-II (43,54). Several studies (34,43,55,56) have demonstrated that it specifically targets ␤-ketoacyl-ACP synthases by covalently attaching to the active site cysteine.…”