Characterization of a Mn-dependent Fructose-1,6-bisphosphate Aldolase in Deinococcus radiodurans |
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Authors: | Yong-Mei Zhang Jong-Kang Liu Mohammad Reza Shouri Tit-Yee Wong |
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Institution: | (1) Department of Microbiology and Molecular Cell Sciences, University of Memphis, Memphis, TN 38152, USA;(2) Department of Biological Sciences, National Sun Yat-sen University, Taiwan, Republic of China;(3) Department of Anatomy & Neurobiology, University of Tennessee Health Science Center, Memphis, TN 38163, USA |
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Abstract: | The key enzyme of the glycolytic pathway of Deinococcus radiodurans, fructose-1,6-bisphosphate aldolase, could be induced independently by glucose and Mn. The enzyme exhibited the characteristics
of the metal-dependent Class II aldolases. Unlike most Class II aldolases, the deinococcal aldolase preferred Mn, not Zn,
as a cofactor. The fbaA gene encoding the deinococcal aldolase was cloned and the protein overproduced in various Escherichia coli expression hosts. However, the overexpressed deinococcal enzyme aggregated and formed inclusion bodies. Dissolving these
inclusion bodies by urea and subsequent purification by nickel affinity chromatography, resulted in a protein fraction that
exhibited aldolase activity only in the presence of Mn. This active aldolase fraction exhibited masses of about 70 kDa and
35 kDa by gel filtration and by SDS gel electrophoresis, respectively, suggesting that the active aldolase was a dimer. |
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Keywords: | class II aldolase Deinococcus radiodurans glycolysis manganese-dependent protein expression |
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