Protein-lipid interactions of human dihydroorotate dehydrogenase and three mutants associated with Miller syndrome.

Abstract

Human dihydroorotate dehydrogenase (DHODH) catalyzes the fourth step of the de novo pyrimidine biosynthesis pathway and uses ubiquinone Q10, a lipophilic molecule located in the inner mitochondrial membrane (IMM), as its co-substrate. DHODH is anchored to the IMM by a single transmembrane helix located at its N-terminus. Nevertheless, how DHODH function is determined by its surrounding membrane environment and protein-lipid interactions, as well as the mechanism by which ubiquinone Q10 accesses the active site of DHODH from within the membrane are still largely unknown. Here, we describe the interaction between wild-type DHODH and three DHODH mutants associated with Miller syndrome and lipids using enzymatic assays, thermal stability assays and Quartz Crystal Microbalance with Dissipation monitoring (QCM-D). Our results provide evidence indicating that the N-terminal part of human DHODH is not only a structural element for mitochondrial import and location of DHODH, but also influences enzymatic activity and utilization of ubiquinone Q10 and ubiquinone analogues in in vitro assays. They also support the role of tetraoleoyl cardiolipin as a lipid interacting with DHODH. Additionally, the results from QCM-D show that the Miller syndrome mutants studied differ in their interactions with supported lipid bilayers compared to wild-type DHODH. These altered interactions add another dimension to the effects of mutations found in Miller syndrome. To the best of our knowledge, this is the first investigation of the protein-lipid interactions of DHODH variants associated with Miller syndrome.

• Authors: Juan Manuel Orozco Rodriguez, Hanna Wacklin-Knecht, Wolfgang Knecht
• Journal: Nucleosides, nucleotides & nucleic acids (Nucleosides Nucleotides Nucleic Acids) Vol. 41 Issue 12 Pg. 1337-1358 ( 2022) ISSN: 1532-2335 [Electronic] United States
• PMID: 35184687 (Publication Type: Journal Article)

Chemical References

• Dihydroorotate Dehydrogenase
• Oxidoreductases Acting on CH-CH Group Donors
• Ubiquinone
• Lipids

Topics

• Humans
• Dihydroorotate Dehydrogenase
• Oxidoreductases Acting on CH-CH Group Donors (genetics, metabolism)
• Ubiquinone (metabolism)
• Lipids