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Functional characte...
Functional characterization of Escherichia coli inorganic pyrophosphatase in zwitterionic buffers
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- Baykov, Alexander A. (författare)
- A. N. Belozersky Institute of Physico-Chemical Biology and School of Chemistry, Moscow State University, Russia
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- Hyytiä, Teppo (författare)
- Department of Chemistry, University of Pennsylvania, USA
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- Turkina, Maria V, 1973- (författare)
- A. N. Belozersky Institute of Physico-Chemical Biology and School of Chemistry, Moscow State University, Russia
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- Efimova, Irina S. (författare)
- A. N. Belozersky Institute of Physico-Chemical Biology and School of Chemistry, Moscow State University, Russia
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- Kasho, Vladimir N. (författare)
- Center for Ulcer Research and Education, Department of Medicine, University of California, Los Angeles, USA
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- Goldman, Adrian (författare)
- Department of Biochemistry University of Turku, Finland; Centre for Biotechnology, University of Turku and Åbo Akademi University, Turku, Finland
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- Cooperman, Barry S. (författare)
- Department of Chemistry, University of Pennsylvania, USA
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- Lahti, Reijo (författare)
- Department of Biochemistry University of Turku, Finland
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A N. Belozersky Institute of Physico-Chemical Biology and School of Chemistry, Moscow State University, Russia Department of Chemistry, University of Pennsylvania, USA (creator_code:org_t)
- 2002-04-21
- 1999
- Engelska.
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Ingår i: European Journal of Biochemistry. - : Wiley-Blackwell Publishing Inc.. - 0014-2956 .- 1432-1033. ; 260:2, s. 308-317
- Relaterad länk:
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https://febs.onlinel...
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https://urn.kb.se/re...
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https://doi.org/10.1...
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Abstract
Ämnesord
Stäng
- Catalysis by Escherichia coli inorganic pyrophosphatase (E-PPase) was found to be strongly modulated by Tris and similar aminoalcoholic buffers used in previous studies of this enzyme. By measuring ligand-binding and catalytic properties of E-PPase in zwitterionic buffers, we found that the previous data markedly underestimate Mg2+-binding affinity for two of the three sites present in E-PPase (3.5- to 16-fold) and the rate constant for substrate (dimagnesium pyrophosphate) binding to monomagnesium enzyme (20- to 40-fold). By contrast, Mg2+-binding and substrate conversion in the enzyme-substrate complex are unaffected by buffer. These data indicate that E-PPase requires in total only three Mg2+ ions per active site for best performance, rather than four, as previously believed. As measured by equilibrium dialysis, Mg2+ binds to 2.5 sites per monomer, supporting the notion that one of the tightly binding sites is located at the trimer–trimer interface. Mg2+ binding to the subunit interface site results in increased hexamer stability with only minor consequences for catalytic activity measured in the zwitterionic buffers, whereas Mg2+ binding to this site accelerates substrate binding up to 16-fold in the presence of Tris. Structural considerations favor the notion that the aminoalcohols bind to the E-PPase active site.
Ämnesord
- NATURVETENSKAP -- Biologi -- Biokemi och molekylärbiologi (hsv//swe)
- NATURAL SCIENCES -- Biological Sciences -- Biochemistry and Molecular Biology (hsv//eng)
Nyckelord
- pyrophosphatase
- magnesium
- Tris
- quaternary structure
- structural modeling
Publikations- och innehållstyp
- ref (ämneskategori)
- art (ämneskategori)
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