(2bi) Extremophile Stress Proteins for Engineering Novel Properties in Living Cells and Biomaterials

Nature has evolved remarkable strategies to survive in adverse environment, as highlighted by the wide diversity of extremophilic organisms that adapt and proliferate under stresses including dehydration, high and low temperatures, or radiation. Throughout my doctoral and postdoctoral trainings, I have investigated distinct classes of extremophilic stress proteins in terms of their native functions as well as potential applications. I envision that understanding stress protein-based extremophilic adaptation would allow the transfer of extremotolerant phenotype to the orthogonal biosystems ranging from prokaryotic and eukaryotic cells to protein-based biomaterials.

Doctoral Research

Advisor: Douglas Clark, University of California, Berkeley

My doctoral project was centered around engineering fibrous chaperone protein called gamma-prefoldin (gPFD) discovered from a hyperthermophilic archaeon Methanocaldococcus jannaschii. Taking advantage of gPFD’s self-assembly and thermostability, I developed enzyme-scaffolded catalytic nanofibers with enhanced enzyme activity and controlled positioning of biomolecules along the filaments through modulating protein-protein interactions. I synthesized gPFD-based hydrogels and demonstrated that properties of an extremophilic protein can be exploited to form functional biomaterials. Lastly, I constructed protein-DNA hybrid nanomaterials using dCas9 as an interfacing component. Thus, I have gained comprehensive protein design skillset for developing functional biomaterials ranging from nanofibers to hydrogels based on extremotolerance-associated protein.

Postdoctoral Research

Advisor: Pamela Silver, Harvard Medical School

In my postdoctoral training, I am broadening the scope of my research into in vivo characterization and engineering of intrinsically disordered proteins (IDPs) from extremophiles. Studying unstructured proteins in living cells complements my doctoral experiences, which focused on in vitro assembly of highly ordered protein nanostructures. In specific, major topics include: 1) Screening of extremotolerance-associated IDPs for the transfer of stress-tolerant phenotypes in human cells, 2) Application of tardigrade IDPs for the stabilization of biological structures, and 3) Engineering synthetic membraneless organelles through liquid-liquid phase separation (LLPS) of tardigrade IDPs. Combined together, these researches comprise a pipeline through which we identify, characterize and transfer useful phenotypes existing in nature.

Teaching Interests

As a graduate student instructor in chemical engineering department, I taught a broad range of courses including chemical engineering core, communication-intensive and elective courses. Based on these experiences, I look forward to teaching core courses in chemical and biomolecular engineering, including but not limited to thermodynamics, reaction kinetics, general / organic chemistry, biochemistry and protein engineering. I am also willing to develop elective courses discussing current research trends in fields such as protein design or biomaterials.

In addition, I have participated in diverse mentorship practices throughout my doctoral and postdoctoral training. As a graduate student, I led a group of undergraduate students and guided their laboratory practices; as I encouraged them to develop their own ideas and actively participate in discussions, many students made meaningful contributions to the research and became co-authors in publications. I also engaged in mentoring high school students through UC Berkeley’s summer internship program, through which I gained passion for nurturing interest in science to the young minds. Currently, I am serving as a voluntary mentor for both Harvard Systems Biology summer internship program and Harvard College Program for Research in Science and Engineering, and I look forward to actively engaging in mentorship and outreach programs in the future.

Selected Publications

• Lim, G. A. Jung, D. J. Glover and D. S. Clark, Enhanced enzyme activity through scaffolding on customizable self-assembling protein filaments, Small, 2019, 15, 1805558. [Selected cover]
• Lim, G. A. Jung, R. Muckom, D. J. Glover and D. S. Clark, Engineering bioorthogonal protein-polymer hybrid hydrogel as a functional protein immobilization platform, Chem. Commun., 2019, 55, 806-809.
• Lim, J. Kim, Y. Kim, D. Xu and D. S. Clark, CRISPR/Cas-directed programmable assembly of multi-enzyme complexes, Chem. Commun., 2020, 56, 4950-4953.
• Lim, D. S. Clark and D. J. Glover, Production of multicomponent protein templates for the positioning and stabilization of enzymes. Methods Mol. Biol., 2020, 2073, 101-115.
• Lim, R.R. Yocum, P.A. Silver and J.C. Way, High spontaneous integration rates of end-modified linear DNAs upon mammalian cell transfection, Sci. Rep., 2023, 13, 6835.
• Lim, S. Ng, D. T. Nguyen, R. T. Quek, J. C. Way and P. A. Silver, Controlling behaviors of intrinsically disordered protein-based synthetic cell compartments using modular oligomerization domains, In preparation.
• Lim, J. C. Way and P. A. Silver, Tardigrade secretory proteins render extracellular desiccation protection, In preparation.

Successful Proposals

• Validation Project Grant ($125k), Wyss Institute for Biologically Inspired Engineering at Harvard University, 2021 – 2023.
• NSF Graduate Research Fellowship (GRFP), 2015 – 2018.