Integral membrane enzymes that catalyze protein lipidation
Covalent attachment of lipids is one of the most prevalent forms of post translational protein modification. Although physico-chemically it amounts to attachment of a mere “hydrophobic stick”, protein lipidation can modulate protein function by altering structure, folding, interaction with other proteins, degradataion and importantly, attachment to membranes. Of the different forms of protein lipidation, the most pervasive form is attachment of a fatty acid to internal cysteines through a thioester. In the cellular milieu, thioesters can be cleaved by thioesterases and thus protein S-acylation, or as it is more commonly known, protein palmitoylation (owing to the prevalence of the attachment of the 16 carbon palmitoyl moiety through this kind of modification) is unique in being the only reversible form of protein lipidation. Protein palmitoylation is catalyzed by members of the DHHC family of eukaryotic integral membrane enzymes so called because of the catalytic Asp-His-His-Cys motif in a cysteine rich domain (CRD) that resides in an intracellular loop. There are 23 DHHC enzymes in humans and several of them have been linked to a number of diseases, especially neuropsychiatric diseases and cancer.
We are interested in the molecular mechanism of DHHC palmitoyltransferases. Very recently we published the first crystal structures of two members of this family (Science, 2018, 359, eaao6326), together with the structure of one of them covalently conjugated to an irreversible inhibitor. these structures have opened up a number of interesting questions that we are currently investigating together with other members in the family.
Molecular mechanism of iron transport
We are focusing on mitochondrial inner membrane transporters that bring iron into mitochondria. Subsequently, the iron is used in the biosynthesis of heme, a central component of the heme in hemoglobin, myoglobin, and cytochromes, and in the biosynthesis of iron-sulfur clusters, important cofactors required for proteins involved in a wide range of cellular activities, namely, electron transport in respiratory chain complexes, regulatory sensing, photosynthesis, and DNA repair. We are currently using heterologous expression to obtain enough purified material for biochemical and biophysical characterization.
Structural and chemical biology approach to design novel antibiotics
Antibiotic-resistant pathogenic bacteria pose a major threat to our healthcare systems. In the face of this challenge, there is a pressing need to identify new targets for combating antibiotic-resistant bacteria and to identify and develop therapeutic leads that can result in clinically useful drugs. Clinically approved antibiotics that are currently in use mostly target bacterial cytosolic enzymes and the ribosome. Integral membrane proteins are, however, a largely uncharted territory for antibiotic development, owing to the difficulty in handling and purification, and importantly, to the lack of structural information. In collaboration with Clifton Barry’s lab, we propose to combine fragment-based drug discovery (FBDD) with high-throughput screening (HTS), together with high-resolution structural analyses, to target integral membrane proteins involved in bacterial cell-envelope biosynthesis and to develop leads for novel antibacterial therapies. In the process, we hope to make fundamental discoveries regarding the mechanistic underpinnings of bacterial cell-envelope biosynthesis.