Enzyme I of the Phosphoenolpyruvate:Sugar Phosphotransferase System: Difference between revisions
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The ~64 kD EI is a homodimers that requires Mg<sup>2+</sup> for phosphorylation by PEP. Each subunit comprises three domains: PEP binds to the C-terminal domain (adopting an α/β barrel fold) and HPr binds to the N-terminal domain (an α-helix bundle). A central domain, tethered to the N-terminal domain by two closely associated linkers and to the C-terminal domain by a long α-helix, contains a phosphorylatable histidine residue (His189). The crystal structure of Escherichia coli EI was obtained by using Mg<sup>2+</sup> and PEP to phosphorylate EI and than adding oxalate (a pyruvate analog). In this structure, His189 is phosphorylated and oriented for in-line phosphotransfer to/from the ligand. Thus, the structure represents an enzyme intermediate just after phosphotransfer from PEP and prior to a conformational transition that brings His189~P in proximity to the phosphoryl group acceptor, His15 of HPr. A model of this conformational transition invokes swiveling around the α-helical linker that disengages the His-domain from the PEP-binding domain. Assuming that HPr binds to the HPr-binding domain as observed by NMR spectroscopy of an EI fragment, a rotation around the two linker segments orients the His-domain relative to the HPr-binding domain so that His189~P and His15 are appropriately stationed for an in-line phosphotransfer reaction. The crystal structure of apo-EI from Staphylococcus carnosus supports this model. | The ~64 kD EI is a homodimers that requires Mg<sup>2+</sup> for phosphorylation by PEP. Each subunit comprises three domains: PEP binds to the C-terminal domain (adopting an α/β barrel fold) and HPr binds to the N-terminal domain (an α-helix bundle). A central domain, tethered to the N-terminal domain by two closely associated linkers and to the C-terminal domain by a long α-helix, contains a phosphorylatable histidine residue (His189). The crystal structure of Escherichia coli EI was obtained by using Mg<sup>2+</sup> and PEP to phosphorylate EI and than adding oxalate (a pyruvate analog). In this structure, His189 is phosphorylated and oriented for in-line phosphotransfer to/from the ligand. Thus, the structure represents an enzyme intermediate just after phosphotransfer from PEP and prior to a conformational transition that brings His189~P in proximity to the phosphoryl group acceptor, His15 of HPr. A model of this conformational transition invokes swiveling around the α-helical linker that disengages the His-domain from the PEP-binding domain. Assuming that HPr binds to the HPr-binding domain as observed by NMR spectroscopy of an EI fragment, a rotation around the two linker segments orients the His-domain relative to the HPr-binding domain so that His189~P and His15 are appropriately stationed for an in-line phosphotransfer reaction. The crystal structure of apo-EI from Staphylococcus carnosus supports this model. | ||
'''''The movie''' depicts the phosphotransfer from PEP to EI and from EI to HPr and the accompanying protein conformational transitions. This is a model based on crystal structure of the intact E. coli EI and the NMR model of the fragment containing the HPr-binding domain and the His-domain in complex with HPr. The HPr-binding domain is colored blue. The PEP-binding domain is colored cyan, the His-domain is colored yellow, and the linker segments that connect the His-domain to the partner domains are colored red. Hpr is colored green. Ligands and the catalytic histidine are depicted in stick models with the atomic color scheme: Carbon – gray, Nitrogen – blue, Oxygen – red, Phosphorous – green, Magnesium – magenta.'' | '''''The movie''' depicts the phosphotransfer from PEP to EI and from EI to HPr and the accompanying protein conformational transitions. This is a model based on crystal structure of the intact E. coli EI and the NMR model of the fragment containing the HPr-binding domain and the His-domain in complex with HPr. The HPr-binding domain is colored blue. The PEP-binding domain is colored cyan, the His-domain is colored yellow, and the linker segments that connect the His-domain to the partner domains are colored red. Hpr is colored green. Ligands and the catalytic histidine are depicted in stick models with the atomic color scheme: Carbon – gray, Nitrogen – blue, Oxygen – red, Phosphorous – green, Magnesium – magenta. The movie was created by Kap Lim and osnat Herzberg'' | ||
== Key References == | == Key References == | ||