1uxh: Difference between revisions
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Revision as of 18:09, 5 November 2007
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LARGE IMPROVEMENT IN THE THERMAL STABILITY OF A TETRAMERIC MALATE DEHYDROGENASE BY SINGLE POINT MUTATIONS AT THE DIMER-DIMER INTERFACE
OverviewOverview
The stability of tetrameric malate dehydrogenase from the green, phototrophic bacterium Chloroflexus aurantiacus (CaMDH) is at least in, part determined by electrostatic interactions at the dimer-dimer, interface. Since previous studies had indicated that the thermal stability, of CaMDH becomes lower with increasing pH, attempts were made to increase, the stability by removal of (excess) negative charge at the dimer-dimer, interface. Mutation of Glu165 to Gln or Lys yielded a dramatic increase in, thermal stability at pH 7.5 (+23.6 -- + 23.9 degrees C increase in, apparent t(m)) and a more moderate increase at pH 4.4 (+4.6 -- + 5.4, degrees C). The drastically increased stability at neutral pH was achieved, without forfeiture of catalytic performance at low temperatures. The, crystal structures of the two mutants showed only minor structural changes, close to the mutated residues, and indicated that the observed stability, effects are solely due to subtle changes in the complex network of, electrostatic interactions in the dimer-dimer interface. Both mutations, reduced the concentration dependency of thermal stability, suggesting that, the oligomeric structure had been reinforced. Interestingly, the two, mutations had similar effects on stability, despite the charge difference, between the introduced side-chains. Together with the loss of, concentration dependency, this may indicate that both E165Q and E165K, stabilize CaMDH to such an extent that disruption of the inter-dimer, electrostatic network around residue 165 no longer limits kinetic thermal, stability.
About this StructureAbout this Structure
1UXH is a Single protein structure of sequence from Chloroflexus aurantiacus with NAD and FMR as ligands. Active as Malate dehydrogenase, with EC number 1.1.1.37 Structure known Active Site: AC1. Full crystallographic information is available from OCA.
ReferenceReference
Large improvement in the thermal stability of a tetrameric malate dehydrogenase by single point mutations at the dimer-dimer interface., Bjork A, Dalhus B, Mantzilas D, Sirevag R, Eijsink VG, J Mol Biol. 2004 Aug 27;341(5):1215-26. PMID:15321717
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