Julnar Al Azzam
Chemistry and Biochemistry
Galia Debelouchina Lab
CBI Appointment Period: 2025-26
|
Elucidating the role of the Y. pestis membrane protein Ail in host-pathogen interactions and outer membrane vesicle formation
Attachment invasion locus, or Ail, is an outer membrane protein native to the plague causing bacterium Yersinia pestis that has been implicated in the immune evasion and host invasion capabilities of the bacterium. Ail has been found to bind the human glycoprotein vitronectin. Vitronectin is involved in downregulating immune responses such as complement-mediated lysis of
infected host cells. When Ail binds vitronectin, it is “shielded” from immune attack. Moreover, as vitronectin binds the surface of epithelial cells, vitronectin-bound Ail can also form a “bridge” between the bacterium and the host cell. The exact interactions between Ail and vitronectin are unknown, therefore, one of the goals of this project is to develop methods for whole-cell NMR of proteins in their native cellular context, where they are most relevant to disease. Moreover, gram-negative bacteria such as Y.pestis and E.coli are known to produce outer membrane vesicles containing genetic material. If Ail is incorporated into these OMV’s, the vesicles may retain the “shielding” and “bridging” capabilities of the whole bacterium and be involved in infection. Another goal of this project is to determine whether Ail plays an active role in OMV formation and is incorporated into the vesicles. OMV’s could provide the possibility for non-live vaccine development that could present antigens in their native context and possess high biodegradability and low acute inflammatory potential. To study these questions, I will use the following tools. Genetic code expansion will be used to incorporate an unnatural amino acid to which several chemical probes can be attached through bio-orthogonal chemistry. For example, I plan to attach a fluorophore, which will allow me to use fluorescence microscopy and perform co-localization experiments with vitronectin. Selective mutation will be used to replace residues in Ail and vitronectin thought to be essential for binding. More detailed structural experiments will be performed in whole cells using solid-state NMR spectroscopy. Finally, I will use the genetic code expansion and bio-orthogonal chemistry strategy to attach gold nanoparticles to Ail. This will allow me to localize the protein by cryo-electron tomography and to investigate its role in OMV formation. To mitigate risks involved in working with Y.pestis, E.coli will
be used for this project as previous experiments have shown that the protein is functional in these cells. Overall, my project will combine chemical biology tools with structural biology methods to investigate fundamental questions related to host-pathogen interactions.
|
Richard Durant
Chemistry and Biochemistry
Michael Burkart Lab
CBI Appointment Period: 2025-26
|
Developing tools for the directed bioengineering of polyketide synthases and non-ribosomal peptide synthetases
My research focuses on developing tools to elucidate the biosynthesis of natural products such as polyketides and nonribosomal peptides. These secondary metabolites represent major classes of bioactive molecules that have historically driven drug discovery and development. Their biosynthetic pathways have attracted great interest since the discovery of the modular architectures of their respective synthases and synthetases, which present opportunities for directed enzyme engineering and combinatorial biosynthesis. Achieving these goals, however, requires reliable manipulation and expression of complex megasynthases which has long posed a significant experimental challenge.
|
Rohan Pillai
Chemistry and Biochemistry
Alexis Komor Lab
CBI Appointment Period: 2025-26
|
HUHe mediated subcellular delivery of structured RNAs
Recent advances in nucleic acid chemistry have led to the development of a range of nucleic acid-based therapies and diagnostic tools. While a variety of RNA bioconjugation strategies have been exploited for cancer and disease treatment, bioconjugation of RNAs to peptide motifs remains an underexplored field of work. While significant work has been performed to enhance the stability and cell-penetrating ability of nucleic acid conjugates, little work has identified strategies to enhance the subcellular delivery of nucleotides toward the organelle of interest. My research explores the in-cellulo conjugation of HUH endonuclease (HUHe) protein-nucleic acid interactions to enhance the
subcellular delivery of structured RNAs towards the nucleus and enable novel delivery of structured RNAs to the mitochondria using CRISPR/Cas genome editing and high-throughput sequencing as an organelle read-out strategy.
|
Sophie Young
Chemistry and Biochemistry
Galia Debelouchina Lab
CBI Appointment Period: 2025-26
|
Developing Selective Fluorescent Sensors for Individual Pathological Calcification Minerals
My project focuses on the development of fluorescent sensors for specific pathological calcification minerals. Ectopic calcification refers to the abnormal accumulation of calcium phosphate minerals in tissues and it is prevalent in chronic and age-related diseases, including cardiovascular disease, age-
related macular degeneration, and Alzheimer’s disease. The specific chemical and molecular mechanisms involved in ectopic calcification are poorly understood. Further, surprisingly little is known about the true mineral composition of calcified deposits or if different minerals have distinct
pathological implications. Current methods to detect and image pathological calcification deposits are unable to distinguish different mineral types. Thus, new tools are needed to determine the presence of specific calcification minerals in tissues and establish their individual roles in the calcification process. To fill this gap, I plan to construct novel protein-based fluorescent sensors that bind and report the presence of specific target minerals. In parallel, to inform sensor development, I will use solid-state nuclear magnetic resonance (NMR) spectroscopy to investigate the mineral-binding interactions of existing small molecule calcification mineral stains.
|