The importance of the strictly conserved, C-terminal glycine residue in phosphoenolpyruvate carboxylase for overall catalysis: mutagenesis and truncation of GLY-961 in the sorghum C4 leaf isoform.

The Journal of Biological Chemistry
Wenxin XuR Chollet

Abstract

Phosphoenolpyruvate carboxylase (PEPC) is a "multifaceted," allosteric enzyme involved in C4 acid metabolism in green plants/microalgae and prokaryotes. Before the elucidation of the three-dimensional structures of maize C4 leaf and Escherichia coli PEPC, our truncation analysis of the sorghum C4 homologue revealed important roles for the enzyme's C-terminal alpha-helix and its appended QNTG961 tetrapeptide in polypeptide stability and overall catalysis, respectively. Collectively, these functional and structural observations implicate the importance of the PEPC C-terminal tetrapeptide for both catalysis and negative allosteric regulation. We have now more finely dissected this element of PEPC structure-function by modification of the absolutely conserved C-terminal glycine of the sorghum C4 isoform by site-specific mutagenesis (G961(A/V/D)) and truncation (DeltaC1/C4). Although the C4 polypeptide failed to accumulate in a PEPC- strain (XH11) of E. coli transformed with the Asp mutant, the other variants were produced at wild-type levels. Although neither of these four mutants displayed an apparent destabilization of the purified PEPC homotetramer, all were compromised catalytically in vivo and in vitro. Functional complementat...Continue Reading

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Citations

Jan 9, 2013·Pharmacogenetics and Genomics·Bernardo B PalmaMichel Kranendonk
Aug 2, 2011·Plant Science : an International Journal of Experimental Plant Biology·Valeria E PerottiFlorencio E Podestá
Aug 2, 2016·Mutation Research. Genetic Toxicology and Environmental Mutagenesis·Bernardo Brito PalmaMichel Kranendonk
Mar 1, 2016·Current Protocols in Plant Biology·Camille Bénard, Yves Gibon

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