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The integral membrane enzyme PagP alternates between two dynamically distinct states.

ABSTRACT: PhoPQ-activated gene P (PagP) is an integral membrane enzyme that transfers the sn-1 palmitate chain from phospholipid to lipopolysaccharide in Gram-negative bacteria. A recent x-ray crystallographic study established that the sn-1 palmitate binds within a long cavity at the center of the PagP beta barrel. The high mobility required to permit substrate entry into the central core of the barrel contrasts with the need to assemble a well defined structure in the peripheral loops, where many key catalytic residues are located. To gain insight into how dynamics relate to the function of PagP, the enzyme was reconstituted into CYFOS-7, a detergent that supports enzymatic activity. Under these conditions, PagP exists in equilibrium between two states, relaxed (R) and tense (T). The kinetics and thermodynamics of the interchange have been investigated by (1)H-(15)N NMR spectroscopy, with Delta H = -10.7 kcal/mol and Delta S = -37.5 cal/mol.K for the R--> T transition. A comparison of chemical shifts between the two states indicates that major structural changes occur in the large extracellular L1 loop and adjacent regions of the beta barrel. In addition to the R,T interconversion, other conformational exchange processes are observed in the R state, showing it to be quite flexible. Thus a picture emerges in which substrate entry is facilitated by the mobility of the R state, whereas the relatively rigid T state adopts a radically different conformation in a region of the protein known to be essential for catalysis. The ability to switch between dynamically distinct states may be a key feature of the catalytic cycle of PagP.

SUBMITTER: Hwang PM

PROVIDER: S-EPMC470724 | biostudies-literature | 2004 Jun

REPOSITORIES: biostudies-literature

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The integral membrane enzyme PagP alternates between two dynamically distinct states.

Hwang Peter M PM Bishop Russell E RE Kay Lewis E LE

Proceedings of the National Academy of Sciences of the United States of America 20040621 26

PhoPQ-activated gene P (PagP) is an integral membrane enzyme that transfers the sn-1 palmitate chain from phospholipid to lipopolysaccharide in Gram-negative bacteria. A recent x-ray crystallographic study established that the sn-1 palmitate binds within a long cavity at the center of the PagP beta barrel. The high mobility required to permit substrate entry into the central core of the barrel contrasts with the need to assemble a well defined structure in the peripheral loops, where many key ca ...[more]

PMID: 15210985

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Project description:Most sensory information destined for the neocortex is relayed through the thalamus, where considerable transformation occurs1,2. One means of transformation involves interactions between excitatory thalamocortical neurons that carry data to the cortex and inhibitory neurons of the thalamic reticular nucleus (TRN) that regulate the flow of those data3-6. Although the importance of the TRN has long been recognised7-9, understanding of its cell types, their organization and their functional properties has lagged behind that of the thalamocortical systems they control. Here we address this by investigating the somatosensory and visual circuits of the TRN in mice. In the somatosensory TRN we observed two groups of genetically defined neurons that are topographically segregated and physiologically distinct, and that connect reciprocally with independent thalamocortical nuclei through dynamically divergent synapses. Calbindin-expressing cells-located in the central core-connect with the ventral posterior nucleus, the primary somatosensory thalamocortical relay. By contrast, somatostatin-expressing cells-which reside along the surrounding edges of the TRN-synapse with the posterior medial thalamic nucleus, a higher-order structure that carries both top-down and bottom-up information10-12. The two TRN cell groups process their inputs in pathway-specific ways. Synapses from the ventral posterior nucleus to central TRN cells transmit rapid excitatory currents that depress deeply during repetitive activity, driving phasic spike output. Synapses from the posterior medial thalamic nucleus to edge TRN cells evoke slower, less depressing excitatory currents that drive more persistent spiking. Differences in the intrinsic physiology of TRN cell types, including state-dependent bursting, contribute to these output dynamics. The processing specializations of these two somatosensory TRN subcircuits therefore appear to be tuned to the signals they carry-a primary central subcircuit tuned to discrete sensory events, and a higher-order edge subcircuit tuned to temporally distributed signals integrated from multiple sources. The structure and function of visual TRN subcircuits closely resemble those of the somatosensory TRN. These results provide insights into how subnetworks of TRN neurons may differentially process distinct classes of thalamic information.

Dataset Information

The integral membrane enzyme PagP alternates between two dynamically distinct states.

Publications

The integral membrane enzyme PagP alternates between two dynamically distinct states.

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