Mukherjee, Arnab - University of California, Santa Barbara - Biomolecular Science and Engineering

个人简介

Ph.D., Chemical Engineering, University of Illinois at Urbana-Champaign B.S./M.S., Biotechnology, Indian Institute of Technology, Madras

研究领域

Engineering genetic reporters for fluorescence imaging in anaerobic systems The great oxygenation event spawned the emergence of oxygen-dependent life on earth as we know it. However, several fascinating and physiologically relevant biosystems continues to thrive in oxygen-free (anaerobic) or low-oxygen (hypoxic) conditions. The poor aqueous solubility of oxygen (7.6 ppm in air-saturated water) coupled with its high reactivity and restricted diffusion in (poorly vascularized) biological tissues can lead to the rapid emergence of anaerobic zones. For instance, large tracts of the mammalian gut are almost entirely anaerobic and have supported the emergence of a complex microbial ecology that profoundly impacts health and disease. Several pathogens exploit hypoxia, frequently associated with soft tissue infections to actively combat the action of antibiotics. Despite the physiological significance of hypoxia, our ability to study cells in low-to-no oxygen conditions has been hindered by a lack of reliable probes that can facilitate biomolecular imaging in these conditions. This is because flagship reporters such as the green fluorescent protein (GFP) and luciferase strictly depend on molecular oxygen to emit light. Consequently, anaerobic biosystems have remained largely “invisible” to the prevalent biomolecular imaging toolkit. To tackle this challenge, we pursue a variety of approaches integrating molecular & metabolic engineering, kinetic modeling, directed evolution, and synthetic biology to engineer bright and variously colored oxygen-independent fluorescent reporters and sensors for anaerobic imaging. Engineering genetic reporters for magnetic resonance imaging Genetically encoded optical reporters based on GFP and luciferase provide one of the most sensitive and selective approaches for imaging molecular level processes in intact cells. Unfortunately, optical techniques provide limited access in biological tissues for the simple reason that photon penetration is restricted to depths of less than one millimeter. Thus, GFP and luciferase-based reporters can not be used to study biological function in the context of intact optically opaque animal models. In contrast to optical techniques, tissue-penetrant modalities such as magnetic resonance imaging (MRI) provide unfettered access in opaque animals. Unfortunately, MRI lacks the molecular precision conventionally reserved for optical reporter genes. Furthermore, relatively few MRI contrast agents have been turned into biochemical sensors for functional imaging. This is a major challenge for biomedical research where animal models are routinely used to study processes such as tumor metastasis and neural function in their important in vivo context. To address this challenge, we engineer biomolecules with unusual properties – for instance, water diffusion and paramagnetism – to develop sensitive and bio-responsive MRI reporter genes for detecting various aspects of physiological function such as tumor gene expression and neural activity.

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Lu, G.J., Farhadi, A., Mukherjee, A., Shapiro, M.G. Proteins, air, and water: reporter genes for ultrasound and magnetic resonance imaging. Current Opinions in Chemical Biology. 45. 57-63. (2018) doi: 10.1016/j.cbpa.2018.02.011 Mukherjee, A., Davis, H., Ramesh, P., Lu, G., Shapiro, M.G. Biomolecular MRI reporters: evolution of new mechanisms. Progress in NMR Spectroscopy. 102-103. 32-42. (2017) doi: 10.1016/j.pnmrs.2017.05.002 Mukherjee, A., Wu, D., Davis, H., Shapiro, M.G. Non-invasive imaging using reporter genes altering cellular water permeability. Nature Communications. 7. 13891. (2016) doi: 10.1038/ncomms13891 Mukherjee, A. and Schroeder, C.M. Flavin-based Fluorescent Proteins: Emerging Paradigms in Biological Imaging. Current Opinions in Biotechnology. 31. 16-23. (2015) doi: 10.1016/j.copbio.2014.07.010 Mukherjee, A., Weyant, K.B., Agrawal, U., Walker, J., Schroeder, C.M. Engineering and Characterization of New LOV-based Fluorescent Reporter Proteins from Chlamydomonas reinhardtii and Vaucheria frigida. ACS Synthetic Biology. (2014) doi: 10.1021/sb500237x Mukherjee, A., Walker, J., Weyant, K.B., Schroeder, C.M. Characterization of Flavin-based Fluorescent Proteins: an Emerging Class of Fluorescent Reporters. PLoS ONE. 8:5. (2013) doi:10.1371/journal.pone.0064753 Mohan, R.‡, Mukherjee, A.‡, Lee, J., Sevgen, S., Schroeder, C.M., Kenis, P.J.A. A Multiplexed Microfluidic Platform for Antibiotic Susceptibility Screening. Biosensors and Bioelectronics. 49:15. 118-125 (2013) doi:10.1016/j.bios.2013.04.046 (‡ denotes equal contribution) Mukherjee, A., Weyant, K.B., Walker, J., Schroeder, C.M. Directed Evolution of Bright Mutants of a Flavin-binding Fluorescent Protein from Pseudomonas putida. Journal of Biol. Engineering. 6:20. (2012) doi: 10.1186/1754-1611-6-20 Schudel, B.R., Tanyeri, M., Mukherjee, A., Schroeder, C.M., Kenis, P.J.A. Multiplexed Detection of Nucleic Acids in a Combinatorial Screening Chip. Lab on a Chip. 11. 1916-23. (2011) doi: 10.1039/C0LC00342E Rajagopala, S.V., Goll, J., Gowda, N.D.D., Sunil, K.C., Titz, B., Mukherjee, A., Mary, S.S., Raviswaran, N., et al. MPI-LIT-A Literature-curated Dataset of Microbial Binary Protein-Protein Interactions. Bioinformatics. 24. 2622-27. (2008) doi: 10.1093/bioinformatics/btn481