Ca2+ stabilizes the semiquinone radical of pyrroloquinoline quinone

2001 ◽  
Vol 357 (3) ◽  
pp. 893-898 ◽  
Author(s):  
Akihiro SATO ◽  
Kazuyoshi TAKAGI ◽  
Kenji KANO ◽  
Nobuo KATO ◽  
Johannis A. DUINE ◽  
...  
Keyword(s):  
Redox Potential ◽  
Active Site ◽  
Continuous Flow ◽  
Glucose Dehydrogenase ◽  
Acinetobacter Calcoaceticus ◽  
Substrate Oxidation ◽  
Pyrroloquinoline Quinone ◽  
Free State ◽  
Semiquinone Radical ◽  
Chemical Mechanisms

Spectroelectrochemical studies were performed on the interaction between Ca2+ and pyrroloquinoline quinone (PQQ) in soluble glucose dehydrogenase (sGDH) and in the free state by applying a mediated continuous-flow column electrolytic spectroelectrochemical technique. The enzyme forms used were holo-sGDH (the holo-form of sGDH from Acinetobacter calcoaceticus) and an incompletely reconstituted form of this, holo-X, in which the PQQ-activating Ca2+ is lacking. The spectroelectrochemical and ESR data clearly demonstrated the generation of the semiquinone radical of PQQ in holo-sGDH and in the free state in the presence of Ca2+. In contrast, in the absence of Ca2+ no semiquinone was observed, either for PQQ in the free state (at pH7.0) or in the enzyme (holo-X). Incorporation of Ca2+ into the active site of holo-X, yielding holo-sGDH, caused not only stabilization of the semiquinone form of PQQ but also a negative shift (of 26.5mV) of the two-electron redox potential, indicating that the effect of Ca2+ is stronger on the oxidized than on the reduced PQQ. Combining these data with the observations on the kinetic and chemical mechanisms, it was concluded that the strong stimulating effect of Ca2+ on the activity of sGDH can be attributed to facilitation of certain kinetic steps, and not to improvement of the thermodynamics of substrate oxidation. The consequences of this conclusion are discussed for the oxidative as well as for the reductive part of the reaction of sGDH.

scholarly journals Reversible thermal inactivation of the quinoprotein glucose dehydrogenase from Acinetobacter calcoaceticus. Ca2+ ions are necessary for re-activation

1989 ◽  
Vol 261 (2) ◽  
pp. 415-421 ◽  
Author(s):  
O Geiger ◽  
H Görisch
Keyword(s):  
Acetic Acid ◽  
Active Site ◽  
Protein Concentration ◽  
Thermal Inactivation ◽  
Glucose Dehydrogenase ◽  
Acinetobacter Calcoaceticus ◽  
Pyrroloquinoline Quinone ◽  
Soluble Form ◽  
Activation Process ◽  
Prosthetic Group

The soluble form of the homogeneous quinoprotein glucose dehydrogenase from Acinetobacter calcoaceticus is reversibly inactivated at temperatures above 35 degrees C. An equilibrium is established between active and denatured enzyme, this depending on the protein concentration and the inactivation temperature used. Upon thermal inactivation the enzyme dissociates into the prosthetic group pyrroloquinoline quinone and the apo form of glucose dehydrogenase. After inactivation at 50 degrees C active enzyme is re-formed again at 25 degrees C. Ca2+ ions are necessary for the re-activation process. The velocity of re-activation depends on the protein concentration, the concentration of the prosthetic group pyrroloquinoline quinone and the Ca2+ concentration. The apo form of glucose dehydrogenase can be isolated, and in the presence of pyrroloquinoline quinone and Ca2+ active holoenzyme is formed. Even though native glucose dehydrogenase is not inactivated in the presence of EDTA or trans-1,2-diaminocyclohexane-NNN'NH-tetra-acetic acid, Ca2+ stabilizes the enzyme against thermal inactivation. Two Ca2+ ions are found per subunit of glucose dehydrogenase. The data suggest that pyrroloquinoline quinone is bound at the active site via a Ca2+ bridge. Mn2+ and Cd2+ can replace Ca2+ in the re-activation mixture.

scholarly journals Pyrroloquinoline Quinone-Dependent Dehydrogenases from Ketogulonicigenium vulgare Catalyze the Direct Conversion of l-Sorbosone to l-Ascorbic Acid

2006 ◽  
Vol 72 (2) ◽  
pp. 1487-1495 ◽  
Author(s):  
Taro Miyazaki ◽  
Teruhide Sugisawa ◽  
Tatsuo Hoshino
Keyword(s):  
Ascorbic Acid ◽  
Glucose Dehydrogenase ◽  
Acinetobacter Calcoaceticus ◽  
Direct Conversion ◽  
Pyrroloquinoline Quinone ◽  
Binding Motif ◽  
Bacterial Cells ◽  
Ketogulonicigenium Vulgare ◽  
Gene Coding ◽  
Prosthetic Groups

ABSTRACT A novel enzyme, l-sorbosone dehydrogenase 1 (SNDH1), which directly converts l-sorbosone to l-ascorbic acid (l-AA), was isolated from Ketogulonicigenium vulgare DSM 4025 and characterized. This enzyme was a homooligomer of 75-kDa subunits containing pyrroloquinoline quinone (PQQ) and heme c as the prosthetic groups. Two isozymes of SNDH, SNDH2 consisting of 75-kDa and 55-kDa subunits and SNDH3 consisting of 55-kDa subunits, were also purified from the bacterium. All of the SNDHs produced l-AA, as well as 2-keto-l-gulonic acid (2KGA), from l-sorbosone, suggesting that tautomerization of l-sorbosone causes the dual conversion by SNDHs. The sndH gene coding for SNDH1 was isolated and analyzed. The N-terminal four-fifths of the SNDH amino acid sequence exhibited 40% identity to the sequence of a soluble quinoprotein glucose dehydrogenase from Acinetobacter calcoaceticus. The C-terminal one-fifth of the sequence exhibited similarity to a c-type cytochrome with a heme-binding motif. A lysate of Escherichia coli cells expressing sndH exhibited SNDH activity in the presence of PQQ and CaCl2. Gene disruption analysis of K. vulgare indicated that all of the SNDH proteins are encoded by the sndH gene. The 55-kDa subunit was derived from the 75-kDa subunit, as indicated by cleavage of the C-terminal domain in the bacterial cells.

scholarly journals Screening of Peptide Ligands for Pyrroloquinoline Quinone Glucose Dehydrogenase Using Antagonistic Template-Based Biopanning

2013 ◽  
Vol 14 (12) ◽  
pp. 23244-23256 ◽  
Author(s):  
Koichi Abe ◽  
Wataru Yoshida ◽  
Kotaro Terada ◽  
Yukiko Yagi-Ishii ◽  
Stefano Ferri ◽  
...  
Keyword(s):  
Glucose Dehydrogenase ◽  
Pyrroloquinoline Quinone ◽  
Peptide Ligands

scholarly journals Active site residues and mechanism of UDP-glucose dehydrogenase

2003 ◽  
Vol 271 (1) ◽  
pp. 14-22 ◽  
Author(s):  
Xue Ge ◽  
Lisa C. Penney ◽  
Ivo Van De Rijn ◽  
Martin E. Tanner
Keyword(s):  
Active Site ◽  
Glucose Dehydrogenase ◽  
Active Site Residues

scholarly journals Identification of Pantoea ananatis gene encoding membrane pyrroloquinoline quinone (PQQ)-dependent glucose dehydrogenase and pqqABCDEF operon essential for PQQ biosynthesis

2011 ◽  
Vol 318 (1) ◽  
pp. 55-60 ◽  
Author(s):  
Irina G. Andreeva ◽  
Lyubov I. Golubeva ◽  
Tatiana M. Kuvaeva ◽  
Evgueni R. Gak ◽  
Joanna I. Katashkina ◽  
...  
Keyword(s):  
Glucose Dehydrogenase ◽  
Pyrroloquinoline Quinone ◽  
Pantoea Ananatis ◽  
Gene Encoding ◽  
Pqq Biosynthesis
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