Raman spectroscopy for monitoring proteome dynamics

Work done in the lab of Prof. Surya Pratap Singh at Integrated Biophotonics Lab, Indian Institute of Technology Dharwad, Karnataka, India.

About author

Jiro Karlo is currently pursuing his 2nd year of PhD in the Department of Bioscience and Bioengineering, IIT Dharwad under the supervision of Dr. Surya Pratap Singh with Institute fellowship (Teaching Assistantship). He has received his Bachelor’s degree in B.Sc. CBZ (Chemistry Botany and Zoology) from H.N.B Garhwal Central University, Uttarakhand. He has done his M.Sc. Life Sciences (Bioinformatics) from Department of Computational Sciences, Central University of Punjab, Bathinda. He has qualified CSIR NET LS/JRF Life Sciences (2020) and GATE XL-Life Sciences (2020). He has published one research article and one review article. He believes in scientific collaboration and discussion to solve problems. He is currently working on the applications of Raman spectroscopy in understanding microbial metabolomics. In his free hours, he loves to play e-sports. His areas of interest are Biophotonics, Raman Spectroscopy, Computational biology, Proteomics and Metabolomics.

Interview

How would you explain your research outcomes to the non-scientific community?

The existing methods to study overall protein-associated changes and turnover in a cell or tissue involve extraction using a destructive process. In this study for the first time, we reported that overall changes in the microbial proteome (turnover, synthesis and degradation) can be identified using Raman spectroscopy. In contrast to existing methods, this approach does not involve any sample processing or extraction. Using non-radioactive isotopes we monitored all the dynamic changes in the protein content of a microbial cell. In addition to being less time-consuming, these findings are expected to provide a solid platform to understand antibiotic resistance, host-pathogen relationship and regulation of bacteria-derived products/metabolites.

The microbial proteome is complex, diverse, and dynamic. It is involved in various biological processes such as host-pathogen interactions, antimicrobial resistance, production of microbial-derived products. Understanding the dynamics of protein and its turnover in the cell’s physiological environment in a stain-free, label free, non-destructive, and non-invasive way is essential for disease diagnostics and therapeutic monitoring. In our study we have successfully demonstrated the potentials of Raman spectroscopy coupled with carbon-13 stable isotope probing to track the de novo global protein synthesis, proteome turnover dynamics and change in proteome level. Our findings suggest Raman stable isotope probing can be an adjunct tool for monitoring proteome level changes in a non-destructive, non-invasive and label-free manner.

How do these findings contribute to your research area?

Stable isotope labelling combined with Raman spectroscopy is an upcoming area. With this study, for the first time, we demonstrated that global dynamic changes in the proteome can be identified using this approach. The findings of the study will provide a solid base for translating this technique in the form of a novel method for assessing dynamic proteome level.

“The findings of the study will provide a solid base for translating this technique in the form of a novel method for assessing dynamic proteome level”

What was the exciting moment during your research?

My ‘eureka’ moment during this work came while performing the inhibition assays involving Kanamycin treatment. When I saw the quantifiable and visible changes in the bands associated with aromatic amino acids and peptide bonds, I felt a great sense of accomplishment.

What do you hope to do next?

I am currently in the second stage of this work which involves tracking the global dynamic changes in the proteome of eukaryotic cells. In the later stage, this work will be performed in animal cells with the help of non-radioactive labelled amino acids.

Where do you seek scientific inspiration from?

Science fiction novels and movies always fascinate me. My biggest inspiration for research comes from the motto that scientific research should not be confined to the laboratory, rather it should be translatable in form of a product or technique that is beneficial to the real world. I always wanted to do research through an interdisciplinary approach combining biological, chemical, and computational sciences. Bio-photonics is a field where one can explore all these fields simultaneously. My parents are one of my biggest strengths. My supervisor is my continuous source of inspiration. He has introduced me to the field of biophotonics and provided me with all guidance, facility, freedom to work, and exposure.

How do you intend to help Indian science improve?

Comparative proteomics is slowly being projected as a tool for various kinds of disease diagnostic and therapeutic monitoring. Cost and extensive sample preprocessing for obtaining the final outcome is a negative factor associated with this approach. With further refinement, the proposed approach can be used as an alternate or adjunct tool that can assist in diagnosis and therapy monitoring.

Reference

Karlo J, Dhillon AK, Siddhanta S, Singh SP. Monitoring of microbial proteome dynamics using Raman stable isotope probing. J Biophotonics. 2022 Dec 17:e202200341. PMID: 36527375. doi: https://doi.org/10.1002/jbio.202200341

Copy Editor

Anjali Mahilkar

Postdoctoral Fellow at University of Michigan

Anjali is an evolutionary biologist working as a postdoctoral researcher in the department of Ecology and Evolutionary Biology at University of Michigan, Ann Arbor. Her research interests mainly lie in understanding the relationship between natural selection, adaptation and drift in deciding the evolutionary fates of biological functions as well as of whole organisms. She has a PhD from IIT Bombay where her research focused on understanding the process of speciation. Anjali also has her Masters from IIT Kanpur and Bachelor’s from NIT Raipur. She also had short work stints at Dr. Reddy’s Institute of Life Science and at a Central University in Chhattisgarh. She is interested in science communication and volunteers for several organizations towards the same. Although not an avid reader, she likes to read fiction in her free time.

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