Enzymatic degradation of cyclic 2,3-diphosphoglycerate to 2,3-diphosphoglycerate in Methanobacterium thermoautotrophicum
2,3-Diphosphoglycerate (2,3-DPG) has been found to be the product of the enzymatic degradation of cyclic 2,3-diphosphoglycerate (cDPG) in the archaebacterium Methanobacterium thermoautotrophicum delta H. Although 2,3-DPG has not previously been detected as a major soluble component of M. thermoautot...
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Published in: | Biochemistry (Easton) Vol. 31; no. 11; pp. 2926 - 2935 |
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American Chemical Society
24-03-1992
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Abstract | 2,3-Diphosphoglycerate (2,3-DPG) has been found to be the product of the enzymatic degradation of cyclic 2,3-diphosphoglycerate (cDPG) in the archaebacterium Methanobacterium thermoautotrophicum delta H. Although 2,3-DPG has not previously been detected as a major soluble component of M. thermoautotrophicum, large pools accumulated at an incubation temperature of 50 degrees C (below the optimum growth temperature of 62 degrees C). Under these conditions, cellular activity was significantly decreased; a return of the culture to the optimum growth temperature restored the 2,3-DPG pool back to original low levels and caused steady-state cDPG levels to increase again. While 13CO2-pulse/12CO2-chase experiments at 50 degrees C showed that the cDPG turned over, the appearance of 2,3-DPG at NMR-visible concentrations required at least 10 h. Production of 2,3-DPG in vivo was prevented by exposure of the cells to O2. The enzyme responsible for this hydrolysis of cDPG was purified by affinity chromatography and appears to be a 33-kDa protein. Activity was detected in the presence of oxygen and was enhanced by a solution of 1 M KCl, 25 mM MgCl2, and dithiothreitol. Both Km and Vmax have been determined at 37 degrees C; kinetics also indicate that in vitro the product, 2,3-DPG, is an inhibitor of cDPG hydrolysis. These findings are discussed in view of a proposed role for cDPG in methanogens. |
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AbstractList | 2,3-Diphosphoglycerate (2,3-DPG) has been found to be the product of the enzymatic degradation of cyclic 2,3-diphosphoglycerate (cDPG) in the archaebacterium Methanobacterium thermoautotrophicum delta H. Although 2,3-DPG has not previously been detected as a major soluble component of M. thermoautotrophicum, large pools accumulated at an incubation temperature of 50 degrees C (below the optimum growth temperature of 62 degrees C). Under these conditions, cellular activity was significantly decreased; a return of the culture to the optimum growth temperature restored the 2,3-DPG pool back to original low levels and caused steady-state cDPG levels to increase again. While 13CO2-pulse/12CO2-chase experiments at 50 degrees C showed that the cDPG turned over, the appearance of 2,3-DPG at NMR-visible concentrations required at least 10 h. Production of 2,3-DPG in vivo was prevented by exposure of the cells to O2. The enzyme responsible for this hydrolysis of cDPG was purified by affinity chromatography and appears to be a 33-kDa protein. Activity was detected in the presence of oxygen and was enhanced by a solution of 1 M KCl, 25 mM MgCl2, and dithiothreitol. Both Km and Vmax have been determined at 37 degrees C; kinetics also indicate that in vitro the product, 2,3-DPG, is an inhibitor of cDPG hydrolysis. These findings are discussed in view of a proposed role for cDPG in methanogens. 2,3-Diphosphoglycerate (2,3-DPG) has been found to be the product of the enzymatic degradation of cyclic 2,3-diphosphoglycerate (cDPG) in the archaebacterium Methanobacterium thermoautotrophicum Delta H. The enzyme responsible for hydrolysis of cDPG was purified by affinity chromatography and appears to be a 33-kDa protein. Activity was detected in the presence of oxygen and was enhanced by a solution of 1 M KCl, 25 mM MgCl sub(2), and dithiothreitol. Both K sub(m) and V sub(max) have been determined at 37 degree C; kinetics also indicate that in vitro the product, 2,3-DPG, is an inhibitor of cDPG hydrolysis. 2,3-Diphosphoglycerate (2,3-DPG) has been found to be the product of the enzymatic degradation of cyclic 2,3-diphosphoglycerate (cDPG) in the archaebacterium Methanobacterium thermoautotrophicum {Delta}H. Although 2,3-DPG has not previously been detected as a major soluble component of M. thermoautotrophicum, large pools accumulated at an incubation temperature of 50C. Under these conditions, cellular activity was significantly decreased; a return of the culture to the optimum growth temperature restored the 2,3-DPG pool back to original low levels and caused steady-state cDPG levels to increase again. While {sup 13}CO{sub 2}-pulse/{sup 12}CO{sub 2}-chase experiments at 50C showed that the cDPG turned over, the appearance of 2,3-DPG at NMR-visible concentrations required at least 10 h. Production of 2,3-DPG in vivo was prevented by exposure of the cells to O{sub 2}. the enzyme responsible for this hydrolysis of cDPG was purified by affinity chromatography and appears to be a 33-kDa protein. Activity was detected in the presence of oxygen and was enhanced by a solution of 1 M KCl, 25 mM MgCl{sub 2}, and dithiothreitol. Both K{sub m} and V{sub max} have been determined at 37C; kinetics also indicate that in vitro the product, 2,3-DPG, is an inhibitor of cDPG hydrolysis. These findings are discussed in view of a proposed role for cDPG in methanogens. |
Author | Sastry, Musti V. Krishna Roberts, Mary F Robertson, Diane E Moynihan, James A |
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Keywords | Temperature Archaeobacteria Enzyme Environmental factor 2,3-Diphosphoglycerate Enzymatic activity Cyclic compound Methanobacteriaceae Enzymatic digestion Bacteria Kinetic parameter Methanobacteriales Methanobacterium thermoautotrophicum |
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Snippet | 2,3-Diphosphoglycerate (2,3-DPG) has been found to be the product of the enzymatic degradation of cyclic 2,3-diphosphoglycerate (cDPG) in the archaebacterium... |
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SubjectTerms | 2,3-Diphosphoglycerate 550601 - Medicine- Unsealed Radionuclides in Diagnostics BACTERIA Bacteriology BIOCHEMICAL REACTION KINETICS BIODEGRADATION Biological and medical sciences CARBOHYDRATES CARBON 12 CARBON 13 CARBON COMPOUNDS CARBON DIOXIDE CARBON ISOTOPES CARBON OXIDES CHALCOGENIDES CHEMICAL REACTIONS CHEMICAL SHIFT Chromatography, Affinity cyclic 2,3-diphosphoglycerate DECOMPOSITION Diphosphoglyceric Acids - metabolism ENZYMATIC HYDROLYSIS ESTERS EVEN-EVEN NUCLEI EVEN-ODD NUCLEI Fundamental and applied biological sciences. Psychology Hydrolases - chemistry Hydrolases - isolation & purification Hydrolases - metabolism HYDROLYSIS ISOTOPES KINETICS LIGHT NUCLEI LIPIDS LYSIS MAGNETIC RESONANCE Magnetic Resonance Spectroscopy Metabolism. Enzymes Methanobacterium - enzymology METHANOGENIC BACTERIA Microbiology MICROORGANISMS NUCLEAR MAGNETIC RESONANCE NUCLEI ORGANIC COMPOUNDS ORGANIC PHOSPHORUS COMPOUNDS OXIDES Oxygen - pharmacology OXYGEN COMPOUNDS PHOSPHOLIPIDS Potassium Chloride - pharmacology RADIOLOGY AND NUCLEAR MEDICINE REACTION KINETICS RESONANCE SOLVOLYSIS Spectrophotometry STABLE ISOTOPES Temperature |
Title | Enzymatic degradation of cyclic 2,3-diphosphoglycerate to 2,3-diphosphoglycerate in Methanobacterium thermoautotrophicum |
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