Research Article: Structural insights into the inhibition properties of archaeon citrate synthase from Metallosphaera sedula

Date Published: February 22, 2019

Publisher: Public Library of Science

Author(s): Seul Hoo Lee, Hyeoncheol Francis Son, Kyung-Jin Kim, Claudio M. Soares.


Metallosphaera sedula is a thermoacidophilic archaeon and has an incomplete TCA/glyoxylate cycle that is used for production of biosynthetic precursors of essential metabolites. Citrate synthase from M. sedula (MsCS) is an enzyme involved in the first step of the incomplete TCA/glyoxylate cycle by converting oxaloacetate and acetyl-CoA into citrate and coenzyme A. To elucidate the inhibition properties of MsCS, we determined its crystal structure at 1.7 Å resolution. Like other Type-I CS, MsCS functions as a dimer and each monomer consists of two distinct domains, a large domain and a small domain. The oxaloacetate binding site locates at the cleft between the two domains, and the active site was more closed upon binding of the oxaloacetate substrate than binding of the citrate product. Interestingly, the inhibition kinetic analysis showed that, unlike other Type-I CSs, MsCS is non-competitively inhibited by NADH. Finally, amino acids and structural comparison of MsCS with other Type-II CSs, which were reported to be non-competitively inhibited by NADH, revealed that MsCS has quite unique NADH binding mode for non-competitive inhibition.

Partial Text

Metallosphaera sedula belongs to the sulfolobaceae family. It is a thermoacidophilic archaea with optimum growth conditions of 73°C and pH 2.0 [1–3]. M. sedula grows chemolithoautotrophically on metal sulfides or molecular hydrogen, and obtains reducing power by biologically catalyzing iron oxidation and metal sulfide oxidation [2]. M. sedula gains access to a carbon source by immobilization of bicarbonate using the 3-hydroxypropionate/4-hydroxybutyrate (3-HP/4-HB) cycle [3–5]. The tricarboxylic acid (TCA)/glyoxylate cycle in this strain is incomplete due to the lack of 2-ketoglutarate dehydrogenase, which converts 2-ketoglutarate to succinyl-CoA, and is used for production of biosynthetic precursors including several amino acids and other essential metabolites in this microorganism [6]. Citrate synthase of M. sedula (MsCS) catalyzes the first step of the incomplete TCA/glyoxylate cycle in this strain.




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