Oxford Molecular Motors

Abstract:

Shewanella oneidensis MR-1 possesses two different stator units to drive flagellar rotation, the Na+-dependent PomAB stator and the H+-driven MotAB stator, the latter possibly acquired by lateral gene transfer. Although either stator can independently drive swimming through liquid, MotAB-driven motors cannot support efficient motility in structured environments or swimming under anaerobic conditions. Using ΔpomAB cells we isolated spontaneous mutants able to move through soft agar. We show that a mutation that alters the structure of the plug domain in MotB affects motor functions and allows cells to swim through media of increased viscosity and under anaerobic conditions. The number and exchange rates of the mutant stator around the rotor were not significantly different from wild-type stators, suggesting that the number of stators engaged is not the cause of increased swimming efficiency. The swimming speeds of planktonic mutant MotAB-driven cells was reduced, and overexpression of some of these stators caused reduced growth rates, implying that mutant stators not engaged with the rotor allow some proton leakage. The results suggest that the mutations in the MotB plug domain alter the proton interactions with the stator ion channel in a way that both increases torque output and allows swimming at decreased pmf values. This article is protected by copyright. All rights reserved.

Abstract:

For the past two decades, the use of genetically fused fluorescent proteins (FPs) has greatly contributed to the study of chemotactic signalling in Escherichia coli including the activation of the response regulator protein CheY and its interaction with the flagellar motor. However, this approach suffers from a number of limitations, both biological and biophysical: for example, not all fusions are fully functional when fused to a bulky FP, which can have a similar molecular weight to its fused counterpart; they may interfere with the native interactions of the protein and the chromophores of FPs have low brightness and photostability and fast photobleaching rates. A recently developed technique for the electroporation of fluorescently labelled proteins in live bacteria has enabled us to bypass these limitations and study the in vivo behaviour of CheY at the single-molecule level. Here we show that purified CheY proteins labelled with organic dyes can be internalized into E. coli cells in controllable concentrations and imaged with video fluorescence microscopy. The use of this approach is illustrated by showing single CheY molecules diffusing within cells and interacting with the sensory clusters and the flagellar motors in real time.

Abstract:

Recent experiments on the bacterial flagellar motor have shown that the structure of this nanomachine, which drives locomotion in a wide range of bacterial species, is more dynamic than previously believed. Specifically, the number of active torque-generating complexes (stators) was shown to vary across applied loads. This finding brings under scrutiny the experimental evidence reporting that limiting (zero-torque) speed is independent of the number of active stators. Here, we propose that, contrary to previous assumptions, the maximum speed of the motor increases as additional stators are recruited. This result arises from our assumption that stators disengage from the motor for a significant portion of their mechanochemical cycles at low loads. We show that this assumption is consistent with current experimental evidence and consolidate our predictions with arguments that a processive motor must have a high duty ratio at high loads.

Abstract:

UNLABELLED:We investigated the characteristics and spatial distribution of cortical bone pores in postmenopausal women with type 2 diabetes (T2D). High porosity in the midcortical and periosteal layers in T2D subjects with fragility fractures suggests that these cortical zones might be particularly susceptible to T2D-induced toxicity and may reflect cortical microangiopathy.

INTRODUCTION:Elevated cortical porosity is regarded as one of the main contributors to the high skeletal fragility in T2D. However, to date, it remains unclear if diabetic cortical porosity results from vascular cortical changes or from an expansion in bone marrow space. Here, we used a novel cortical laminar analysis technique to investigate the characteristics and spatial radial distribution of cortical pores in a T2D group with prior history of fragility fractures (DMFx, assigned high-risk group) and a fracture-free T2D group (DM, assigned low-risk group) and to compare their results to non-diabetic controls with (Fx) and without fragility fractures (Co).

METHODS:Eighty postmenopausal women (n = 20/group) underwent high-resolution peripheral quantitative computed tomography (HR-pQCT) of the distal tibia and radius. Cortical bone was divided into three layers of equal width including an endosteal, midcortical, and periosteal layer. Within each layer, total pore area (TPA), total pore number (TPN), and average pore area (APA) were calculated. Statistical analysis employed Mann-Whitney tests and ANOVA with post hoc tests.

RESULTS:Compared to the DM group, DMFx subjects exhibited +90 to +365 % elevated global porosity (p = 0.001). Cortical laminar analysis revealed that this increased porosity was for both skeletal sites confined to the midcortical layer, followed by the periosteal layer (midcortical +1327 % TPA, p ≤ 0.001, periosteal +634 % TPA, p = 0.002), and was associated in both layers and skeletal sites with high TPN (+430 % TPN, p < 0.001) and high APA (+71.5 % APA, p < 0.001).

CONCLUSION:High porosity in the midcortical and periosteal layers in the high-risk T2D group suggests that these cortical zones might be particularly susceptible to T2D-induced toxicity and may reflect cortical microangiopathy.

Abstract:

Cosmology has made enormous progress through studies of the cosmic microwave background, however the subtle signals being now sought such as B-mode polarisation due to primordial gravitational waves are increasingly hard to disentangle from residual Galactic foregrounds in the derived CMB maps. We revisit our finding that on large angular scales there are traces of the nearby old supernova remnant Loop I in the WMAP 9-year map of the CMB and confirm this with the new SMICA map from the Planck satellite.