Crystal structure of FabZ-ACP complex reveals a dynamic seesaw-like catalytic mechanism of dehydratase in fatty acid biosynthesis

Abstract

Fatty acid biosynthesis (FAS) is a vital process in cells. Fatty acids are essential for cell assembly and cellular metabolism. Abnormal FAS directly correlates with cell growth delay and human diseases, such as metabolic syndromes and various cancers. The FAS system utilizes an acyl carrier protein (ACP) as a transporter to stabilize and shuttle the growing fatty acid chain throughout enzymatic modules for stepwise catalysis. Studying the interactions between enzymatic modules and ACP is, therefore, critical for understanding the biological function of the FAS system. However, the information remains unclear due to the high flexibility of ACP and its weak interaction with enzymatic modules. We present here a 2.55 Å crystal structure of type II FAS dehydratase FabZ in complex with holo-ACP, which exhibits a highly symmetrical FabZ hexamer-ACP3 stoichiometry with each ACP binding to a FabZ dimer subunit. Further structural analysis, together with biophysical and computational results, reveals a novel dynamic seesaw-like ACP binding and catalysis mechanism for the dehydratase module in the FAS system, which is regulated by a critical gatekeeper residue (Tyr100 in FabZ) that manipulates the movements of the β-sheet layer. These findings improve the general understanding of the dehydration process in the FAS system and will potentially facilitate drug and therapeutic design for diseases associated with abnormalities in FAS.

Structural and dynamical rationale for fatty acid unsaturation in Escherichia coli

“…S5). The AcpP-interacting surface of E. coli FabZ is strictly analogous to the AcpP-interacting surface of H. pylori FabZ (20). However, in contrast to our finding that the E. coli FabZ hexamer can interact productively with six AcpPs, a see-saw mechanism was proposed for the H. pylori FabZ in which only three AcpPs simultaneously engage the hexamer.…”

“…3 and SI Appendix, Table S2). The FabZ hexamer is a trimer of FabA-like dimers, consistent with structures of other bacterial FabZ enzymes (13,20,(25)(26)(27). In the hexamer, an AcpP is cross-linked to the catalytic His54 of each FabZ monomer and contacts both subunits of the FabZ dimer but makes no contacts with the other two "dimers" or with any other AcpP ( Figs.…”

“…However, the H. pylori FabZ 6 -AcpP 3 stoichiometry in the crystal structure is more simply explained by crystal lattice contacts that block the three unoccupied AcpP sites. Previous surface plasmon resonance (SPR) characterization of the H. pylori FabZ-AcpP interaction indicated a single K d of 120 nM (23), in contrast to the three weaker binding events deduced from MST data (20). In our SEC-MALS analysis of the E. coli FabZ-AcpP interaction, higher-order complexes existed with an excess of either apoor holo-AcpP, and a 20-fold excess AcpP fully saturated the Mocr-FabZ hexamer ( Fig.…”

“…1C), the second step in the transformation, remains speculative (12,17,19). Recently, the crystal structure of a complex of phosphopantetheinylated AcpP (holo-AcpP) and FabZ from Helicobacter pylori was described (20), but the ACP lacked an acyl group, limiting conclusions regarding the distinct activities and substrate preferences of FabA and FabZ.…”

The structural organization of substrate loading in iterative polyketide synthases

“…By contrast, symmetric large-scale conformational transitions that require partial refolding of conserved domains have been visualized by EM analysis of distinct functional states of the modPKS PikAIII on the basis of a divergent condensing region architecture 12 , 26 . Functional asymmetry in the interaction of ACP with dimeric enzymatic domains has previously been suggested in the form of a seesaw-type conformational coupling for a bacterial hexameric dehydratase (FabZ) involved in fatty acid biosynthesis 34 that binds ACPs in a 6:3 ratio. However, a stoichiometrically ACP-crosslinked structure of the bacterial dimeric dehydratase FabA demonstrates that asymmetry is not a general feature in related dehydratases even though this structure also revealed increased disorder for the second ACP, as indicated by the crystallographic B factors 25 .…”