The Histidine Phosphocarrier Kinase/Phosphorylase from Bacillus Subtilis Is an Oligomer in Solution with a High Thermal Stability
Por:
Neira, J, Camara-Artigas, A, Hernandez-Cifre, J and Ortore, M
Publicada:
1 mar 2021
Resumen:
The histidine phosphocarrier protein (HPr) kinase/phosphorylase (HPrK/P) modulates the phosphorylation state of the HPr protein, and it is involved in the use of carbon sources by Gram-positive bacteria. Its X-ray structure, as concluded from crystals of proteins from several species, is a hexamer; however, there are no studies about its conformational stability, and how its structure is modified by the pH. We have embarked on the conformational characterization of HPrK/P of Bacillus subtilis (bsHPrK/P) in solution by using several spectroscopic (namely, fluorescence and circular dichroism (CD)) and biophysical techniques (namely, small-angle X-ray-scattering (SAXS) and dynamic light-scattering (DLS)). bsHPrK/P was mainly a hexamer in solution at pH 7.0, in the presence of phosphate. The protein had a high conformational stability, with an apparent thermal denaturation midpoint of similar to 70 degrees C, at pH 7.0, as monitored by fluorescence and CD. The protein was very pH-sensitive, precipitated between pH 3.5 and 6.5; below pH 3.5, it had a molten-globule-like conformation; and it acquired a native-like structure in a narrow pH range (between pH 7.0 and 8.0). Guanidinium hydrochloride (GdmCl) denaturation occurred through an oligomeric intermediate. On the other hand, urea denaturation occurred as a single transition, in the range of concentrations between 1.8 and 18 mu M, as detected by far-UV CD and fluorescence.
Filiaciones:
:
Univ Miguel Hernandez, IDIBE, Alicante 03202, Spain
Univ Zaragoza, Inst Biocomputac & Fis Sistemas Complejos, Joint Units IQFR CSIC BIFI, Zaragoza 50009, Spain
Univ Zaragoza, GBsC CSIC BIFI, Zaragoza 50009, Spain
Camara-Artigas, A:
Univ Almeria ceiA3, Dept Quim & Fis, Res Ctr CIAIMBITAL, Almeria 04120, Spain
Hernandez-Cifre, J:
Univ Murcia, Dept Quim Fis, Fac Quim, Campus Espinardo, Murcia 30100, Spain
Ortore, M:
Univ Politecn Marche, Dipartimento DiSVA, Via Brecce Bianche, I-60131 Ancona, Italy
Green Published, gold
|