1e0u
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(New page: 200px<br /> <applet load="1e0u" size="450" color="white" frame="true" align="right" spinBox="true" caption="1e0u, resolution 2.8Å" /> '''STRUCTURE R271L MUTA...)
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Revision as of 06:52, 18 November 2007
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STRUCTURE R271L MUTANT OF E. COLI PYRUVATE KINASE
Overview
Pyruvate kinase (PK) is critical for the regulation of the glycolytic, pathway. The regulatory properties of Escherichia coli were investigated, by mutating six charged residues involved in interdomain salt bridges, (Arg(271), Arg(292), Asp(297), and Lys(413)) and in the binding of the, allosteric activator (Lys(382) and Arg(431)). Arg(271) and Lys(413) are, located at the interface between A and C domains within one subunit. The, R271L and K413Q mutant enzymes exhibit altered kinetic properties. In, K413Q, there is partial enzyme activation, whereas R271L is characterized, by a bias toward the T-state in the allosteric equilibrium. In the, T-state, Arg(292) and Asp(297) form an intersubunit salt bridge. The, mutants R292D and D297R are totally inactive. The crystal structure of, R292D reveals that the mutant enzyme retains the T-state quaternary, structure. However, the mutation induces a reorganization of the interface, with the creation of a network of interactions similar to that observed in, the crystal structures of R-state yeast and M1 PK proteins. Furthermore, in the R292D structure, two loops that are part of the active site are, disordered. The K382Q and R431E mutations were designed to probe the, binding site for fructose 1, 6-bisphosphate, the allosteric activator., R431E exhibits only slight changes in the regulatory properties., Conversely, K382Q displays a highly altered responsiveness to the, activator, suggesting that Lys(382) is involved in both activator binding, and allosteric transition mechanism. Taken together, these results support, the notion that domain interfaces are critical for the allosteric, transition. They couple changes in the tertiary and quaternary structures, to alterations in the geometry of the fructose 1, 6-bisphosphate and, substrate binding sites. These site-directed mutagenesis data are, discussed in the light of the molecular basis for the hereditary, nonspherocytic hemolytic anemia, which is caused by mutations in human, erythrocyte PK gene.
About this Structure
1E0U is a Single protein structure of sequence from Escherichia coli with SO4 as ligand. The following page contains interesting information on the relation of 1E0U with [The Glycolytic Enzymes]. Active as Pyruvate kinase, with EC number 2.7.1.40 Full crystallographic information is available from OCA.
Reference
The allosteric regulation of pyruvate kinase., Valentini G, Chiarelli L, Fortin R, Speranza ML, Galizzi A, Mattevi A, J Biol Chem. 2000 Jun 16;275(24):18145-52. PMID:10751408
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