4.A.1 The PTS Glucose-Glucoside (Glc) Family
The Glc family includes porters specific for glucose, glucosamine, N-acetylglucosamine and a large variety of α- and β-glucosides. However, not all β-glucoside PTS porters are in this class, as the PTS porter first described (as the cellobiose (Cel) β-glucoside porter) is the diacetylchitobiose porter in the Lac family (TC #4.A.3). The IIA, IIB and IIC domains of all of the group translocators listed below are demonstrably homologous. These porters (the IIC domains) show limited sequence similarity with and are homologous to members of the Fru family (TC #4.A.2) and with (but less so) members of the Lac family (TC #4.A.3)and the Gut (glucitol) family (TC# 4.A.4). The IIC domains of the glucose (4.A.1.1) and glucoside (4.A.1.2) subfamilies are nearly as distant from each other as they are from the Fru, Mtl and Lac families. The Gut family is more distant due to an internal rearrangement relative to members of the other families mintioned. As is true of other members of the PTS-GFL superfamily, the IIC domains of these permeases probably have a uniform 10 TMS topology (McCoy et al. 2016; Vastermark and Saier 2016). Butanol toxicity in Clostridium acetobutylicum results in destruction of the PTS, thereby preventing glucose transport and phosphorylation (Gao et al. 2021).
Several of the PTS porters in the Glc family lack their own IIA domains and instead use the glucose IIA protein (IIAglc or Crr). Most of these porters have the B and C domains linked together in a single polypeptide chain. A cysteyl residue in the IIB domain is phosphorylated by direct phosphoryl transfer from IIAglc(his~P) or one of its homologues. Those porters which lack a IIA domain include the maltose (Mal), arbutin-salicin-cellobiose (ASC), trehalose (Tre), putative glucoside (Glv) and sucrose (Scr) porters of E. coli. Most, but not all Scr porters of other bacteria also lack their own IIA domains.
BglF consists of a transmembrane domain, which in addition to TMSs, contains a large cytoplasmic loop. According to Yagur-Kroll et al., 2009, this loop, connecting TMSI to TMSII, contains regions that alternate between facing-in and facing-out states and creates the sugar translocation channel. Yagur-Kroll et al., 2009 demonstrated spatial proximity between positions at the center of the big loop and the phosphorylation site, suggesting that these two regions come together to execute sugar phosphotransfer.
The three-dimensional structures of the IIA and IIB domains of the E. coli glucose porter have been elucidated. IIAglc has a complex β-sandwich structure while IIBglc is a split αβ-sandwich with a topology unrelated to the split αβ-sandwich structure of HPr. Some bacteria have many PTS transport systems belonging to different families. For example, the solventogenic Clostridium acetobutylicum ATCC 824 has 13 altogether with 6 in the Glc family, 2 in the Fru family, 2 in the Lac family, 1 in the Gat family and 2 in the Man family. However, Clostridium beijerinckii has 43 phosphotransferases (Mitchell 2015).
Structures of an N,N′-diacetylchitobiose EIIC transporter bcChbC (7) and a maltose EIIC transporter bcMalT, both from Bacillus cereus,
have been reported. bcChbC and bcMalT share 19% sequence identity and
50% similarity, and both have the same structural fold with almost
all of the secondary structural elements conserved (Ren et al. 2018).
Both proteins are homodimers, and each protomer has 10 transmembrane sequenes
(TMSs 1-10), two reentrant loops (HP1-2), and two amphipathic
helices (AH1-2). These structural elements fold into two distinctive
structural domains. The dimerization domain (also referred to as the
interface domain), which consists of TMSs 1-5 and AH1, forms an expansive
dimer interface. The substrate-binding domain (also referred to as the
transport domain), which is composed of TMSs 6-10 and two reentrant loops
(HP1-2), contains the sugar-binding site. In both structures, the sugar
substrate is coordinated by residues from TMSs 6 and 7, HP1, and HP2. The two domains are bridged by an amphipathic helix (AH2).
The bcChbC and bcMalT structures represent different conformations required to complete a transport cycle.
Based on the location of the substrate-binding site, bcChbC is in an
inward-facing conformation, while bcMalT is in an outward-facing
conformation (Ren et al. 2018).
When the two structures are aligned by their dimerization domains, the substrate-binding domain can carry the substrate across
the membrane by a rigid-body motion. A similar elevator-type transport
mechanism has been reported in a number of secondary solute
transporters including amino acid transporters (EAAT1 (TC# 2/A/23/2/1) and GltPh (TC# 2.A.23.1.5)), bile acid transporters (ASBT; 2.A.28.1.2), proton sodium exchangers (NhaA: TC# 2.A.33.1.1), concentrative nucleotide transporters (CNTNW; TC# 2.A.41.2.6), and citrate transporters (vcINDY and SeCitS).
These transporters have different structural folds, and yet they all
transport substrates from one side of the cell membrane to the other by
rigid-body motions of a substrate-binding domain.
Although
by comparing the inward-facing bcChbC structure and the outward-facing
bcMalT structure one can postulate that the glucose superfamily of EIICs
have an elevator-type mechanism of transport, we need to visualize both
conformations in the same transporter to reveal the
conformational changes. To achieve this, Ren et al. 2018 first generated a structural
model of bcMalT in an inward-facing conformation by collective
variable-based steered molecular dynamics (CVSMD) simulation using the bcChbC structure as a guide.
During the simulation, the interface domain was kept static, and the
substrate-binding domain was steered toward the inward-facing
conformation. Since the substrate-binding domain moves relative to the
interface domain, distance changes between the two domains were expected.
Indeed, the CVSMD model showed that residues that were far away from each
other in the outward-facing structure became closer, for example,
residues T280 and D55 and residues N284 and E54. The pairs of residues predicted to become closer to
each other can be cross-linked by a mercury ion when mutated to
cysteine residues and thus provide an experimental validation to the
CVSMD model and the elevator-type mechanism of transport.
Ren et al. 2018 solved the crystal structure of bcMalT cross-linked in
an inward-facing conformation. The structure provided direct
experimental evidence that the substrate-binding domain can undergo a
rigid-body rotation toward the intracellular side. The structure also
showed conformational changes in other regions of the transporter that
accommodate the rigid-body movement of the substrate-binding domain.
This family belongs to the: PTS-GFL Superfamily.
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