Protein structure: Get into the groove(-tunnel)

Lipidomics Gateway (28 September 2011) [doi:10.1038/lipidmaps.2011.28]

Insights from the crystal structure of the thioesterase domain of fatty acid synthase bound by a polyunsaturated fatty acyl adduct might offer unique opportunities for the development of effective fatty acid synthase inhibitors.

Fatty acid synthase (FAS) is required for the de novosynthesis of long-chain fatty acids, predominantly palmitate, but is overexpressed, coincident with an increase in de novo fatty acid synthesis, in cancer and several other pathophysiological processes. Conversely, long-chain polyunsaturated fatty acids (PUFAs) show anti-proliferative effects, often in conjunction with inhibition of FAS activity. Now, a report by Zhang et al. of the crystal structure of the thioesterase (TE) domain of FAS with a polyunsaturated fatty acyl adduct provides insight into how PUFAs might confer their beneficial effects.

The TE domain is actually one of seven different catalytic domains in the homodimeric multifunctional FAS. Located at the carboxyl terminus, it both determines the length of the fatty acyl chain and releases the fatty acid. In their study, Zhang et al. determined the X-ray structure of the TE domain of human FAS (hFAS) covalently modified by a methylfluorophosphonate head group attached to a γ-linolenyl acyl tail (methyl γ-linolenyl fluorophosphonate (MGLFP)) at 1.48 Å resolution, revealing several binding features. Consistent with the known reactivity of MGLFP, the phosphonate head group phosphorylated a serine in the active site of TE, abrogating thioesterase activity. Unexpectedly, however, the 18-carbon polyunsaturated γ-linolenyl chain becomes buried within a long contiguous groove-tunnel site, forming complementary interactions with amino acid residues within it. This groove-tunnel is formed by a 'gatekeeper helix' or 'helix flap', an amphipathic α-helix that emerges as a consequence of ligand binding, and has not previously been observed in TE structural studies.

Because the γ-linolenyl tail bound so precisely to the TE domain, Zhang et al. surmised that other PUFAs might behave similarly, and therefore investigated the influence of three omega-6 linolenic acids — the dietary PUFAs γ-linolenic acid (GLA) and α-linolenic acid (ALA), and the GLA elongation product, dihomo-γ-linolenic acid (DGLA) — on TE domain esterase activity. DGLA inhibited TE domain activity much more effectively than GLA or ALA, although inhibition of the activity of intact hFAS by DGLA was slightly less effective compared with the isolated domain.

Next, the authors extended their studies to assess the effects of DGLA on de novo fatty acid synthesis in cells. Incubation of 3T3-L1 preadipocytes with DGLA reduced the incorporation of [¹³C]-sodium acetate into [¹³C]-palmitate by ~50%, indicating that DGLA inhibited palmitate biosynthesis. In human breast cancer cell lines, the ethyl ester of DGLA inhibited the activity of FAS, resulting in a decrease in fatty acid biosynthesis. Furthermore, the DGLA ethyl ester decreased the viability of the breast cancer cells compared with noncancerous cells, although the authors acknowledge that the precise mechanism responsible for this decrease requires further investigation.

The authors also recognize that, although their structural studies have uncovered a molecular binding mechanism for γ-linolenyl acyl chain and, by extension from the results of modelling experiments, DGLA, precisely how the TE domain binds other acyl chains, such as palmitoyl, requires a more complete atomic-level understanding. Nevertheless, the finding that polyunsaturated fatty acyl chains induce the formation of a groove-tunnel site within the active site of TE provides what the authors describe as “a tantalizing unique avenue of approach in developing FAS TE inhibitors with greater specificity and potency.”

Katrin Legg

References:

ORIGINAL RESEARCH PAPER

1. Zhang, W. et al. Crystal structure of FAS thioesterase domain with polyunsaturated fatty acyl adduct and inhibition by dihomo-γ-linolenic acid.
   Proc. Nat. Acad. Sci. USA (2011). doi:10.1073/pnas.1112334108

FURTHER READING

1. Menendez, J. A. & Lupu, R. Fatty acid synthase and the lipogenic phenotype in cancer pathogenesis.
   Nature Rev. Cancer 7, 763-777 (2007). doi:10.1038/nrc2222