Crystal structure of type-III geranylgeranyl pyrophosphate synthase from Saccharomyces cerevisiae and the mechanism of product chain length determination

Geranylgeranyl pyrophosphate synthase (GGPPs) catalyzes a condensation reaction of farnesyl pyrophosphate with isopentenyl pyrophosphate to generate C₂₀ geranylgeranyl pyrophosphate, which is a precursor for carotenoids, chlorophylls, geranylgeranylated proteins, and archaeal ether-linked lipid. For short-chain trans-prenyltransferases that synthesize C₁₀-C₂₅ products, bulky amino acid residues generally occupy the fourth or fifth position upstream from the first DDXXD motif to block further elongation of the final products. However, the short-chain type-III GGPPs in eukaryotes lack any large amino acid at these positions. In this study, the first structure of type-III GGPPs from Saccharomyces cerevisiae has been determined to 1.98 Å resolution. The structure is composed entirely of 15 α-helices joined by connecting loops and is arranged with α-helices around a large central cavity. Distinct from other known structures of trans-prenyltransferases, the N-terminal 17 amino acids (9-amino acid helix A and the following loop) of this GGPPs protrude from the helix core into the other subunit and contribute to the tight dimer formation. Deletion of the first 9 or 17 amino acids caused the dissociation of dimer into monomer, and the Δ(1-17) mutant showed abolished enzyme activity. In each subunit, an elongated hydrophobic crevice surrounded by D, F, G, H, and I α-helices contains two DDXXD motifs at the top for substrate binding with one Mg ²⁺ coordinated by Asp⁷⁵, Asp⁷⁹, and four water molecules. It is sealed at the bottom with three large residues of Tyr ¹⁰⁷, Phe¹⁰⁸, and His¹³⁹. Compared with the major product C₃₀ synthesized by mutant H139A, the products generated by mutant Y107A and F108A are predominantly C₄₀ and C₃₀, respectively, suggesting the most important role of Tyr¹⁰⁷ in determining the product chain length.