Enzyme I of the Phosphoenolpyruvate:Sugar Phosphotransferase System: Difference between revisions
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[[Image:Pts.jpg|left|thumb|The PTS|400px]]<br> | [[Image:Pts.jpg|left|thumb|The PTS|400px]]<br> | ||
Bacteria evolved a unique mechanism to import many carbohydrates | Bacteria evolved a unique mechanism to import many carbohydrates, the '''phosphoenolpyruvate:sugar phosphotransferase system''' (PTS). The PTS synchronizes the transport and phosphorylation of the sugar (group translocation), engaging several proteins in a five steps phosphorylation cascade. With some variations, the PTS comprises three proteins: In the cytoplasm, PEP phosphorylates '''Enzyme I''' (EI), which then transfers the phosphoryl group to the histidine phosphocarrier protein, HPr. From HPr, the phosphoryl group is transferred to various sugar-specific membrane associated transporters (EnzII), each comprising two cytoplasmic domains, EIIA and EIIB, and an integral membrane domain EIIC. Within EII, EIIA accepts the phosphoryl group from HPr and donates it to EIIB, whereupon EIIC mediates sugar translocation with EIIB providing the phosphoryl group. In addition to controlling sugar translocation, the phosphorylation state of PTS proteins is also associated with regulation of metabolic pathways and signaling in bacterial cells.<br> | ||
[[Image:Enzidimer.jpg|left|thumb|Enzyme I dimer|250px]] | [[Image:Enzidimer.jpg|left|thumb|Enzyme I dimer|250px]] | ||
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. | ||