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Engineering a temperature sensitive tobacco etch virus protease

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Molecular engineering study demonstrates rapid conditional proteolysis in synthetic cell systems, highlighting new strategies for cellular control.

To engineer a conditionally controlled, temperature-sensitive tobacco etch virus protease capable of rapid and specific target cleavage.
Generated temperature-sensitive variants of tobacco etch virus protease (tsTEVp) using N-terminal truncations.
Evaluated cleavage efficiency and kinetics following thermal induction at 30°C compared to baseline activity at 37°C.
Assayed downstream functional outputs in cell models, including protein translocation, targeted protein degradation, and Ca2+-mediated cellular blebbing.
N-terminally truncated tsTEVp exhibited robust proteolytic cleavage within 4 hours at 30°C while remaining largely dormant at 37°C.
Thermal activation of tsTEVp successfully drove diverse cellular responses, including targeted protein translocation, degradation, and calcium-mediated membrane blebbing.

Wong et al. (2017) studied this question.

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Molecular engineering study demonstrates rapid conditional proteolysis in synthetic cell systems, highlighting new strategies for cellular control.