No abstract

In Bacteroidetes, one of the dominant phyla of the mammalian gut, active uptake of large nutrients across the outer membrane is mediated by SusCD protein complexes via a “pedal bin” transport mechanism. However, many features of SusCD function in glycan uptake remain unclear, including ligand binding, the role of the SusD lid and the size limit for substrate transport. Here we characterise the β2,6 fructo-oligosaccharide (FOS) importing SusCD from Bacteroides thetaiotaomicron (Bt1762-Bt1763) to shed light on SusCD function. Co-crystal structures reveal residues involved in glycan recognition and suggest that the large binding cavity can accommodate several substrate molecules, each up to ~2.5 kDa in size, a finding supported by native mass spectrometry and isothermal titration calorimetry. Mutational studies in vivo provide functional insights into the key structural features of the SusCD apparatus and cryo-EM of the intact dimeric SusCD complex reveals several distinct states of the transporter, directly visualising the dynamics of the pedal bin transport mechanism.

No abstract

Caseinolytic protein, ClpC is a general stress protein which belongs to the heat shock protein HSP100 family of molecular chaperones. Some of the Clp group proteins have been identified as having a role in the pathogenesis of many bacteria. The Mycobacterium tuberculosis genome demonstrates the presence of a ClpC homolog, ClpC1. M. tuberculosis ClpC1 is an 848‐amino acid protein, has two repeat sequences at its N‐terminus and contains all the determinants to be classified as a member of the HSP100 family. In this study, we overexpressed, purified and functionally characterized M. tuberculosis ClpC1. Recombinant M. tuberculosis ClpC1 showed an inherent ATPase activity, and prevented protein aggregation. Furthermore, to investigate the contribution made by the N‐terminal repeats of ClpC1 to its functional activity, two deletion variants, ClpC1Δ1 and ClpC1Δ2, lacking N‐terminal repeat I and N‐terminal repeat I along with the linker between N‐terminal repeats I and II, respectively were generated. Neither deletion affected the ATPase activity. However, ClpC1Δ1 was structurally altered, less stable and was unable to prevent protein aggregation. Compared with wild‐type protein, ClpC1Δ2 was more active in preventing protein aggregation and displayed higher ATPase activity at high pH values and temperatures. The study demonstrates that M. tuberculosis ClpC1 manifests chaperone activity in the absence of any adaptor protein and only one of the two N‐terminal repeats is sufficient for the chaperone activity. Also, an exposed repeat II makes the protein more stable and functionally more active.

Transient receptor potential (TRP) ion channels are eukaryotic polymodal sensors that function as molecular cellular signal integrators. TRP family members sense and are modulated by a wide array of inputs, including temperature, pressure, pH, voltage, chemicals, lipids, and other proteins. These inputs induce signal transduction events mediated by nonselective cation passage through TRP channels. In this review, we focus on the thermosensitive TRP channels and highlight the emerging view that these channels play a variety of significant roles in physiology and pathophysiology in addition to sensory biology. We attempt to use this viewpoint as a framework to understand the complexity and controversy of TRP channel modulation and ultimately suggest that the complex functional behavior arises inherently because this class of protein is exquisitely sensitive to many diverse and distinct signal inputs. To illustrate this idea, we primarily focus on TRP channel thermosensing. We also offer a structural, biochemical, biophysical, and computational perspective that may help to bring more coherence and consensus in understanding the function of this important class of proteins.