Contribution of Special Structural Features to High Thermal Stability of a Cold-active Transglutaminase
YI Zhang, Chen Li, Timothy Geary, Benjamin Kofi Simpson
Journal of Agriculture and Food Chemistry
A cold-active transglutaminase (TGase, EC 126.96.36.199) that catalyzes the reaction of protein glutamine + protein lysine ↔ protein with γ-glutamyl-ε-lysine cross-link + NH3 at low temperatures was reported previously. This study verified the thermal stability of the TGase from 0–80 oC. Fluorescence and CD spectra studies confirmed tertiary structural damage at 40 oC, α-helix reduction at 60 oC, and refolding during cooling to 20 oC. The TGase sequence was obtained by transcriptomics and used to build its structure. Its catalytic triad was Cys333–His403–Asp426 and its catalytic process was inferred from the model. Molecular dynamics simulation illustrated that its cold-activity resulted from its flexible active site, while high thermostability was conferred by overall rigid structure, a large amount of stable Val, Lys, and strong electrostatic interactions at the N- and C- terminals. This study fills gaps in the correlation of conformational changes with stability and activity of TGase.
Circular dichroism, Secondary structure, Thermal stability, Protein folding, Biochemistry