2-Succinyl-6-hydroxy-2,4-cyclohexadiene-1-carboxylate synthase (EC 4.2.99.20), also known as SHCHC synthase is encoded by the menH gene in Escherichia coli and functions in the synthesis of vitamin K.^[1] The specific step in the synthetic pathway that SHCHC synthase catalyzes is the conversion of 5-enolpyruvoyl-6-hydroxy-2-succinylcyclohex-3-ene-1-carboxylate to (1R,6R)-6-hydroxy-2-succinylcyclohexa-2,4-diene-1-carboxylate and pyruvate.^[2]

Vitamin K is a fat soluble vitamin known to aid in blood clotting. It is recommended that all newborns receive an injection of vitamin K in order to prevent excessive bleeding of the brain after birth. There are two major forms of vitamin K that occur naturally. Phylloquinone, also known as K₁, is synthesized by plants and is the major form of vitamin K in the diet. Menaquinone, K₂, includes a range of forms that are synthesized by bacteria in the gut.^[3]

Vitamin K is synthesized from the molecule chorismate in a nine step conversion process. SHCHC synthase catalyzes the third step in the process.^[4]

The crystal structure of the MenH enzyme in E.coli (SHCHC synthase) exists as a complex of three protein molecules shown in the diagram. SHCHC synthase forms an alpha/beta hydrolase fold with a central set of seven parallel beta sheets surrounded by alpha helices on both sides. A cap of five alpha helixes serves to enclose the active site.^[5] The enzyme exists in an open form until it binds the substrate, when it morphs into a closed form with an active catalytic triad.^[6]

Energetic analysis shows that SHCHC synthase has a low energetic burden for catalytic activity.^[1] This means the enzyme is more prone to mutation and is one of the most diverse enzymes in the vitamin K synthetic pathway.^[7] Only fifteen amino acid residues are absolutely conserved across mutations of the enzyme.^[7]

The active site contains a catalytic triad of syrine, histine and arginine, which is conserved across all mutants and is proposed to initiate the reaction.^[1] The triad residues are located at Ser86, Asp210, and His232.^[5] This triad is proposed to catalyze a proton extraction which triggers a transfer of electrons leading to the elimination of pyruvate and formation of SHCHC.^[6] Originally, it was proposed that the transition state was stabilized by a nontraditional oxyanion hole. Now a traditional oxyanion hole is favored, but not definitive.^[5]

SHCHC synthase is unaffected by traditional cofactors such as divalent metal ions and EDTA.^[1] The enzyme is fairly specific and only acts on SEPHCHC and close derivatives.^[2]

MenH (SHCHC synthase) was previously thought to be a thioesterase involved in hydrolyzing DHNA-CoA in a later step of menaquinone synthesis. In 2008, it was determined that MenH has poor catalytic activity toward palmitoyl-CoA, casting doubt on its role as a thioesterase.^[1] Direct analysis confirmed that MenH is unable to hydrolyze DHNA-CoA.^[1] In 2009, it was proposed that a dedicated hotdog fold thioesterase would be needed to catalyze the hydrolysis of DHNA-CoA.^[8] A protein was identified in 2013 that could fit this role.^[9]