Recombinant Expression of 15N/13C-Labeled Ubiquitin

Human ubiquitin G76C was expressed in Escherichia coli strain BL21 (DE3) in LB media (Nacalai Tesque) or M9 minimal media containing 99% ¹⁵N-labeled ammonium chloride (Cambridge Isotope Laboratories); for the triple-resonance NMR experiments, the M9 medium also contained 99% U-¹³C-labeled D-glucose (Cambridge Isotope Laboratories). The ubiquitin mutant was purified as described previously^{5 (link)}. Protein purity was checked by SDS-PAGE.

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Morimoto D., Walinda E., Shinke M., Sugase K, & Shirakawa M. (2018). Isolation and characterization of a minimal building block of polyubiquitin fibrils. Scientific Reports, 8, 2711.

Publication 2018

15N-labeled ammonium chloride

Ammonium chloride Escherichia coli Glucose Human ubiquitin Isotope Protein Resonance Sds page Strain Ubiquitin

Corresponding Organization : Kyoto University

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Variable analysis

independent variables

Expression host: Escherichia coli strain BL21 (DE3)
Culture media: LB media (Nacalai Tesque) or M9 minimal media containing 99% 15N-labeled ammonium chloride (Cambridge Isotope Laboratories)

dependent variables

Protein purity: Checked by SDS-PAGE

control variables

Protein: Human ubiquitin G76C mutant
Purification: As described previously (link provided)

controls

No positive or negative controls were explicitly mentioned in the provided information.

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