Structural insight into the PTS sugar transporter EIIC

Abstract

Background: The enzyme IIC (EIIC) component of the phosphotransferase system (PTS) is responsible for selectively transporting sugar molecules across the inner bacterial membrane. This is accomplished in parallel with phosphorylation of the sugar, which prevents efflux of the sugar back across the membrane. This process is a key part of an extensive signaling network that allows bacteria to efficiently utilize preferred carbohydrate sources.

Scope of review: The goal of this review is to examine the current understanding of the structural features of the EIIC and how it mediates concentrative, selective sugar transport. The crystal structure of an N,N'-diacetylchitobiose transporter is used as a structural template for the glucose superfamily of PTS transporters.

Major conclusions: Comparison of protein sequences in context with the known EIIC structure suggests that members of the glucose superfamily of PTS transporters may exhibit variations in topology. Despite these differences, a conserved histidine and glutamate appear to have roles shared across the superfamily in sugar binding and phosphorylation. In the proposed transport model, a rigid body motion between two structural domains and movement of an intracellular loop provide the substrate binding site with alternating access, and reveal a surface required for interaction with the phosphotransfer protein responsible for catalysis.

General significance: The structural and functional data discussed here give a preliminary understanding of how transport in EIIC is achieved. However, given the great sequence diversity between varying glucose-superfamily PTS transporters and lack of data on conformational changes needed for transport, additional structures of other members and conformations are still required. This article is part of a Special Issue entitled: Structural biochemistry and biophysics of membrane proteins.

Keywords: ChbC; Enzyme IIC; Membrane protein; Phosphotransferase system; Sugar transport; Transporter.

Figure 1

Organization of the PTS. A. Flow diagram illustrating sequence of phosphorylation events leading to the addition of phosphate (yellow) to the EIIC transporter-bound sugar molecule (red). B. The EIIC domain can be translated individually or as a multi-domain protein containing EIIB and/or EIIA.

Figure 2

The bcChbC structure. A. The two protomers of the bcChbC dimer are colored green and cyan. Each monomer contains a molecule of N,N’-diacetylchitobiose. Each protomer contains two domains: an N-terminal dimerization domain made up of TM1 through TM5 and a C-terminal domain in which transport and phosphorylation occur containing TM6-10. B. The N,N’-diacetylchitobiose binding site. Interacting residues are colored yellow and originate from TM6, TM7, HP1b, TM8, and the reentrant loop containing HP2.

Figure 3

Topology diagram of the bcChbC monomer. The bcChbC protomer contains ten transmembrane helices and two reentrant loops. The protein is oriented with the periplasmic side on top, and helices are indicated with green rectangles. Beta sheets are indicated with yellow arrows. The N-terminal oligomerization domain and the C-terminal transport domain are separated by a periplasmic amphipathic helix (AH2).

Figure 4

Aligment of the EIIC domain of four glucose superfamily members based on transmembrane helix prediction by TMHMM. Predicted transmembrane helices are in red. Transmembrane helices observed in the bcChbC crystal structure are in yellow, reentrant loop helices are in green, and E334 and H250 in bcChbC are blue. Predicted helices from gene fusion and cysteine labeling experiments are in black boxes. The conserved region identified by Nguyen et al. is highlighted with a pink box.

Figure 5

Predicted model of the bcChbC bcChbB complex. The electrostatic surfaces of A. bcChbC and B. a homology model of bcChbB are complementary. C. The predicted bcChbC (green) bcChbB (cyan) complex generated by manual docking. The cysteine attached phosphate is positioned directly below the O6 oxygen of maltose (magenta).

Figure 6

Model of the (A) inward open ChbC monomer crystal structure and (B) predicted ChbC monomer outward open state involving a rigid body rotation and upwards translation of the transport domain (TM6-10, green) relative to the oligomerization domain (TM1-5, cyan).

Figure 7

Putative bcChbC gates controlling sugar entry and exit from the transporter. Residues that align in sequence with mutations in PtsG that affect transport are colored yellow and labeled according to the PtsG sequence. Protomer 1 is green, protomer 2 is cyan, N,N’-diacetylchitobiose is magenta, and E334 and H250 are shown in blue.