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J. Biol. Chem., Vol. 281, Issue 21, 14991-15000, May 26, 2006

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Crystal Structure of Type-III Geranylgeranyl Pyrophosphate Synthase from Saccharomyces cerevisiae and the Mechanism of Product Chain Length Determination^*

Tao-Hsin Chang, Rey-Ting Guo, Tzu-Ping Ko^¶, Andrew H.-J. Wang^¶1, and Po-Huang Liang^¶2

From the Institute of Biochemical Sciences, National Taiwan University, Taipei 106, Taiwan, the Taiwan International Graduate Program, Academia Sinica, Taipei 115, Taiwan, and the ^¶Institute of Biological Chemistry, Academia Sinica, Taipei 115, Taiwan

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 -helices joined by connecting loops and is arranged with -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 (1-17) mutant showed abolished enzyme activity. In each subunit, an elongated hydrophobic crevice surrounded by D, F, G, H, and I -helices contains two DDXXD motifs at the top for substrate binding with one Mg²⁺ coordinated by Asp⁷⁵, Asp⁷⁹, and four water molecules. It is sealed at the bottom with three large residues of Tyr¹⁰⁷, 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.

Received for publication, December 2, 2005 , and in revised form, February 24, 2006.

The atomic coordinates and structure factors (code 2DH4) have been deposited in the Protein Data Bank, Research Collaboratory for Structural Bioinformatics, Rutgers University, New Brunswick, NJ (http://www.rcsb.org/).

^* This work was supported by National Science Council Grant NSC94-2311-B-001-034 and a grant from Academia Sinica (to P.-H. L.). The Biological Crystallography Facilities (Taiwan Beamline BL12B2 at SPring-8) were supported by the National Science Council. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ To whom correspondence may be addressed: Institute of Biological Chemistry, Academia Sinica, 128 Academia Rd., Taipei 115, Taiwan. Tel.: 886-2-2785-5696 (ext. 5010); Fax: 886-2-2788-9759; E-mail: ahjwang{at}gate.sinica.edu.tw. ² To whom correspondence may be addressed: Institute of Biological Chemistry, Academia Sinica, 128 Academia Rd., Taipei 115, Taiwan. Tel.: 886-2-2785-5696 (ext. 6070); Fax: 886-2-2788-9759; E-mail: phliang{at}gate.sinica.edu.tw.

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