A mild skeletal phenotype with overlapping features of Miller syndrome and functional characterisation of two new variants of human dihydroorotate dehydrogenase.

Dihydroorotate dehydrogenase (DHODH) catalyzes the fourth enzymatic reaction of the pyrimidine biosynthesis pathway. Miller syndrome, also known as postaxial acrofacial dysostosis, is caused by biallelic pathogenic variants in . We present a patient with a relatively mild skeletal phenotype carrying a novel variant of unknown significance in : c.

Recombinant Protein Production Using the Baculovirus Expression Vector System (BEVS).

The baculovirus expression vector system (BEVS) is one of the most popular eukaryotic systems for recombinant protein production. The focus of our protein production platform is the provision of recombinant proteins for research use, where generally only small quantities are required, in the range of tens of micrograms to a few hundred milligrams. Here, we present methods that reflect our standard operating procedures and setup to be able to frequently, and often repeatedly, produce many different types of proteins.

New Insights into the Interaction of Class II Dihydroorotate Dehydrogenases with Ubiquinone in Lipid Bilayers as a Function of Lipid Composition.

The fourth enzymatic reaction in the de novo pyrimidine biosynthesis, the oxidation of dihydroorotate to orotate, is catalyzed by dihydroorotate dehydrogenase (DHODH). Enzymes belonging to the DHODH Class II are membrane-bound proteins that use ubiquinones as their electron acceptors. We have designed this study to understand the interaction of an N-terminally truncated human DHODH (Δ29DHODH) and the DHODH from (DHODH) with ubiquinone (Q) in supported lipid membranes using neutron reflectometry (NR).

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

Human dihydroorotate dehydrogenase (DHODH) catalyzes the fourth step of the pyrimidine biosynthesis pathway and uses ubiquinone Q, 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 Q accesses the active site of DHODH from within the membrane are still largely unknown.

Protein production, kinetic and biophysical characterization of three human dihydroorotate dehydrogenase mutants associated with Miller syndrome.

Miller syndrome is a rare Mendelian disorder caused by mutations in the gene encoding human dihydroorotate dehydrogenase (DHODH). Human DHODH, a Class II DHODH, is an integral protein of the inner mitochondrial membrane (IMM) catalyzing the fourth step of the pyrimidine biosynthesis pathway. Here we present a summary of the state of knowledge regarding Miller syndrome in the absence of any current review on the topic.

Preparation of human dihydroorotate dehydrogenase for interaction studies with lipid bilayers.

Human dihydroorotate dehydrogenase (DHODH) is an integral protein of the inner mitochondrial membrane (IMM) that catalyzes the fourth step of the pyrimidine biosynthesis and is functionally connected to the respiratory chain via its lipophilic co-substrate, ubiquinone Q. DHODH is the target for drugs approved for the treatment of rheumatoid arthritis and multiple sclerosis, and mutations in its sequence have been identified as the cause of Miller syndrome, a rare genetic disorder. The N-terminus of DHODH consists of a signal peptide for mitochondrial import (MS), a transmembrane domain (TM), followed by a microdomain which interacts with the lipids of the IMM and has been proposed to form the binding site for ubiquinone Q.