NMR characterization of a 264-residue hyperthermostable endo-beta-1,3-glucanase

Biochemical and Biophysical Research Communications
Johannes H IppelCarlo P M van Mierlo

Abstract

Insight into the hyperthermostable endo-beta-1,3-glucanase pfLamA from Pyrococcus furiosus is obtained by using NMR spectroscopy. pfLamA functions optimally at 104 degrees C and recently the X-ray structure of pfLamA has been obtained at 20 degrees C, a temperature at which the enzyme is inactive. In this study, near-complete (>99%) NMR assignments are presented of chemical shifts of pfLamA in presence and absence of calcium at 62 degrees C, a temperature at which the enzyme is biologically active. The protein contains calcium and the effects of calcium on the protein are assessed. Calcium binding results in relatively small chemical shift changes in a region distant from the active site of pfLamA and thus causes only minor conformational modifications. Removal of calcium does not significantly alter the denaturation temperature of pfLamA, implying that calcium does not stabilize the enzyme against global unfolding. The data obtained form the basis for elucidation of the molecular origins involved in conformational stability and biological activity of hyperthermophilic endo-beta-1,3-glucanases at extreme temperatures.

References

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Citations

May 15, 2013·Applied Microbiology and Biotechnology·Agata PrzybyszWillem J H van Berkel
Jul 17, 2013·Magnetic Resonance in Medicine : Official Journal of the Society of Magnetic Resonance in Medicine·Dung Minh HoangYoussef Zaim Wadghiri

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