Structure Report for: 5SYN

Labby, K.J., Stuckey, J.A., Meagher, J.L., Won, S.J., Martin, B.R.

Title: Molecular Mechanism for Isoform-Selective Inhibition of Acyl Protein Thioesterases 1 and 2 (APT1 and APT2)
Won SJ, Davda D, Labby KJ, Hwang SY, Pricer R, Majmudar JD, Armacost KA, Rodriguez LA, Rodriguez CL and Martin BR <7 more author(s)> Won SJ, Davda D, Labby KJ, Hwang SY, Pricer R, Majmudar JD, Armacost KA, Rodriguez LA, Rodriguez CL, Chong FS, Torossian KA, Palakurthi J, Hur ES, Meagher JL, Brooks CL, 3rd, Stuckey JA, Martin BR (- 7)
Ref: ACS Chemical Biology, 11:3374, 2016 : PubMed
Abstract


ESTHER: Won_2016_ACS.Chem.Biol_11_3374
PubMedSearch: Won 2016 ACS.Chem.Biol 11 3374
PubMedID: 27748579
Gene_locus related to this paper: human-LYPLA2
Inhibitor(s) related to this paper: ML349, ML348

Abstract

Post-translational S-palmitoylation directs the trafficking and membrane localization of hundreds of cellular proteins, often involving a coordinated palmitoylation cycle that requires both protein acyl transferases (PATs) and acyl protein thioesterases (APTs) to actively redistribute S-palmitoylated proteins toward different cellular membrane compartments. This process is necessary for the trafficking and oncogenic signaling of S-palmitoylated Ras isoforms, and potentially many peripheral membrane proteins. The depalmitoylating enzymes APT1 and APT2 are separately conserved in all vertebrates, suggesting unique functional roles for each enzyme. The recent discovery of the APT isoform-selective inhibitors ML348 and ML349 has opened new possibilities to probe the function of each enzyme, yet it remains unclear how each inhibitor achieves orthogonal inhibition. Herein, we report the high-resolution structure of human APT2 in complex with ML349 (1.64 A), as well as the complementary structure of human APT1 bound to ML348 (1.55 A). Although the overall peptide backbone structures are nearly identical, each inhibitor adopts a distinct conformation within each active site. In APT1, the trifluoromethyl group of ML348 is positioned above the catalytic triad, but in APT2, the sulfonyl group of ML349 forms hydrogen bonds with active site resident waters to indirectly engage the catalytic triad and oxyanion hole. Reciprocal mutagenesis and activity profiling revealed several differing residues surrounding the active site that serve as critical gatekeepers for isoform accessibility and dynamics. Structural and biochemical analysis suggests the inhibitors occupy a putative acyl-binding region, establishing the mechanism for isoform-specific inhibition, hydrolysis of acyl substrates, and structural orthogonality important for future probe development.