Abstract
Adenosylcobalamin-dependent diol dehydratase is one of essential components of carboxysome-like polyhedral bodies. It exists as a heterohexamer (αβγ)2, and its activity is recovered in a precipitant fraction of Klebsiella oxytoca and overexpressing Escherichia coli cells. Limited proteolysis of the enzyme with trypsin converted the enzyme into a highly soluble form without loss of enzyme activity. The N-terminal amino acid sequencing of the enzyme thus solubilized indicated that the N-terminal 20 and 16 amino acid residues had been removed from the β and γ subunits, respectively. Mutant enzymes with the same N-terminal truncations of either or both of the β and γ subunits were expressed on a high level in E. coli cells. All the mutant enzymes obtained were expressed in a soluble, active form. These results indicate that the N-terminal regions of the β and γ subunits lower the solubility of diol dehydratase. The mutant enzyme with the N-terminal truncations of both β and γ subunits was essentially indistinguishable in catalytic properties from recombinant wild-type enzyme or the enzyme purified from K. oxytoca in a soluble form.
Original language | English |
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Pages (from-to) | 455-462 |
Number of pages | 8 |
Journal | Bioscience, Biotechnology and Biochemistry |
Volume | 69 |
Issue number | 3 |
DOIs | |
Publication status | Published - Mar 2005 |
Keywords
- Adenosylcobalamin
- Coenzyme B
- Diol dehydratase
- Glycerol dehydratase
- Solubility of enzyme
ASJC Scopus subject areas
- Biotechnology
- Analytical Chemistry
- Biochemistry
- Applied Microbiology and Biotechnology
- Molecular Biology
- Organic Chemistry