TY - JOUR
T1 - The chondroitin sulfate/dermatan sulfate 4-O-endosulfatase from marine bacterium Vibrio sp FC509 is a dimeric species: Biophysical characterization of an endosulfatase
AU - Neira, José L.
AU - Medina-Carmona, Encarnación
AU - Hernández-Cifre, José G.
AU - Montoliu-Gaya, Laia
AU - Cámara-Artigás, Ana
AU - Seffouh, Ilham
AU - Gonnet, Florence
AU - Daniel, Régis
AU - Villegas, Sandra
AU - de la Torre, José García
AU - Pey, Angel L.
AU - Li, Fuchuan
PY - 2016/12/1
Y1 - 2016/12/1
N2 - © 2016 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM) Sulfatases catalyze hydrolysis of sulfate groups. They have a key role in regulating the sulfation states that determine the function of several scaffold molecules. Currently, there are no studies of the conformational stability of endosulfatases. In this work, we describe the structural features and conformational stability of a 4-O-endosulfatase (EndoV) from a marine bacterium, which removes specifically the 4-O-sulfate from chondroitin sulfate/dermatan sulfate. For that purpose, we have used several biophysical techniques, namely, fluorescence, circular dichroism (CD), FTIR spectroscopy, analytical ultracentrifugation (AUC), differential scanning calorimetry (DSC), mass spectrometry (MS), dynamic light scattering (DLS) and size exclusion chromatography (SEC). The protein was a dimer with an elongated shape. EndoV acquired a native-like structure in a narrow pH range (7.0–9.0); it is within this range where the protein shows the maximum of enzymatic activity. The dimerization did not involve the presence of disulphide-bridges as suggested by AUC, SEC and DLS experiments in the presence of β-mercaptoethanol (β-ME). EndoV secondary structure is formed by a mixture of α and β-sheet topology, as judged by deconvolution of CD and FTIR spectra. Thermal and chemical denaturations showed irreversibility and the former indicates that protein did not unfold completely during heating.
AB - © 2016 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM) Sulfatases catalyze hydrolysis of sulfate groups. They have a key role in regulating the sulfation states that determine the function of several scaffold molecules. Currently, there are no studies of the conformational stability of endosulfatases. In this work, we describe the structural features and conformational stability of a 4-O-endosulfatase (EndoV) from a marine bacterium, which removes specifically the 4-O-sulfate from chondroitin sulfate/dermatan sulfate. For that purpose, we have used several biophysical techniques, namely, fluorescence, circular dichroism (CD), FTIR spectroscopy, analytical ultracentrifugation (AUC), differential scanning calorimetry (DSC), mass spectrometry (MS), dynamic light scattering (DLS) and size exclusion chromatography (SEC). The protein was a dimer with an elongated shape. EndoV acquired a native-like structure in a narrow pH range (7.0–9.0); it is within this range where the protein shows the maximum of enzymatic activity. The dimerization did not involve the presence of disulphide-bridges as suggested by AUC, SEC and DLS experiments in the presence of β-mercaptoethanol (β-ME). EndoV secondary structure is formed by a mixture of α and β-sheet topology, as judged by deconvolution of CD and FTIR spectra. Thermal and chemical denaturations showed irreversibility and the former indicates that protein did not unfold completely during heating.
KW - Circular dichroism
KW - Differential scanning calorimetry
KW - Fluorescence
KW - Irreversibility
KW - Mass spectrometry
KW - Protein stability
U2 - 10.1016/j.biochi.2016.09.015
DO - 10.1016/j.biochi.2016.09.015
M3 - Article
VL - 131
SP - 85
EP - 95
ER -
