Macromolecular condensation organizes nucleolar sub-phases to set up a pH gradient.

Nucleoli are multicomponent condensates defined by coexisting sub-phases. We identified distinct intrinsically disordered regions (IDRs), including acidic (D/E) tracts and K-blocks interspersed by E-rich regions, as defining features of nucleolar proteins. We show that the localization preferences of nucleolar proteins are determined by their IDRs and the types of RNA or DNA binding domains they encompass. In vitro reconstitutions and studies in cells showed how condensation, which combines binding and complex coacervation of nucleolar components, contributes to nucleolar organization. D/E tracts of nucleolar proteins contribute to lowering the pH of co-condensates formed with nucleolar RNAs in vitro. In cells, this sets up a pH gradient between nucleoli and the nucleoplasm. By contrast, juxta-nucleolar bodies, which have different macromolecular compositions, featuring protein IDRs with very different charge profiles, have pH values that are equivalent to or higher than the nucleoplasm. Our findings show that distinct compositional specificities generate distinct physicochemical properties for condensates.

Structure and Mechanisms of F-Type ATP Synthases.

F1Fo ATP synthases produce most of the ATP in the cell. F-type ATP synthases have been investigated for more than 50 years, but a full understanding of their molecular mechanisms has become possible only with the recent structures of complete, functionally competent complexes determined by electron cryo-microscopy (cryo-EM). High-resolution cryo-EM structures offer a wealth of unexpected new insights. The catalytic F1 head rotates with the central γ-subunit for the first part of each ATP-generating power stroke. Joint rotation is enabled by subunit δ/OSCP acting as a flexible hinge between F1 and the peripheral stalk. Subunit a conducts protons to and from the c-ring rotor through two conserved aqueous channels. The channels are separated by ∼6 Šin the hydrophobic core of Fo, resulting in a strong local field that generates torque to drive rotary catalysis in F1. The structure of the chloroplast F1Fo complex explains how ATPase activity is turned off at night by a redox switch. Structures of mitochondrial ATP synthase dimers indicate how they shape the inner membrane cristae. The new cryo-EM structures complete our picture of the ATP synthases and reveal the unique mechanism by which they transform an electrochemical membrane potential into biologically useful chemical energy.

Determinants, maintenance, and function of organellar pH.

The protonation state of soluble and membrane-associated macromolecules dictates their charge, conformation, and functional activity. In addition, protons (H+ or their equivalents) partake in numerous metabolic reactions and serve as a source of electrochemical energy to drive the transmembrane transport of both organic and inorganic substrates. Stringent regulation of the intracellular pH is therefore paramount to homeostasis. Although the regulation of the cytosolic pH has been studied extensively, our understanding of the determinants of the H+ concentration ([H+]) of intracellular organelles has developed more slowly, limited by their small size and inaccessibility. Recently, however, targeting of molecular probes to the organellar lumen together with advances in genomic, proteomic, and electrophysiological techniques have led to the identification and characterization of unique pumps, channels, and transporters responsible for the establishment and maintenance of intraorganellar pH. These developments and their implications for cellular function in health and disease are the subject of this review.

Translation arrest cancellation of VemP, a secretion monitor in Vibrio, is regulated by multiple cis- and trans-factors, including SecY.

VemP is a secretory protein in the Vibrio species that monitors cellular protein-transport activity through its translation arrest, allowing expression of the downstream secD2-secF2 genes in the same operon, which encode components of the protein translocation machinery. When cellular protein-transport function is fully active, secD2/F2 expression remains repressed as VemP translation arrest is canceled immediately. The VemP arrest-cancellation occurs on the SecY/E/G translocon in a late stage in the translocation process and requires both trans-factors, SecD/F and PpiD/YfgM, and a cis-element, Arg-85 in VemP; however, the detailed molecular mechanism remains elusive. This study aimed to elucidate how VemP passing through SecY specifically monitors SecD/F function. Genetic and biochemical studies showed that SecY is involved in the VemP arrest-cancellation and that the arrested VemP is stably associated with a specific site in the protein-conducting pore of SecY. VemP-Bla reporter analyses revealed that a short hydrophobic segment adjacent to Arg-85 plays a critical role in the regulated arrest-cancellation with its hydrophobicity correlating with the stability of the VemP arrest. We identified Gln-65 and Pro-67 in VemP as novel elements important for the regulation. We propose a model for the regulation of the VemP arrest cancellation by multiple cis-elements and trans-factors with different roles.

Discovery and structural characterization of the D-box, a conserved TonB motif that couples an inner-membrane motor to outer-membrane transport.

Gram-negative bacteria use TonB-dependent transport to take up nutrients from the external environment, employing the Ton complex to import a variety of nutrients that are either scarce or too large to cross the outer membrane unaided. The Ton complex contains an inner-membrane motor (ExbBD) that generates force, as well as nutrient-specific transport proteins on the outer membrane. These two components are coupled by TonB, which transmits the force from the inner to the outer membrane. TonB contains an N-terminus anchored in the inner membrane, a C-terminal domain that binds the outer-membrane transporter, and a proline-rich linker connecting the two. While much is known about the interaction between TonB and outer-membrane transporters, the critical interface between TonB and ExbBD is less well understood. Here, we identify a conserved motif within TonB that we term the D-box, which serves as an attachment point for ExbD. We characterize the interaction between ExbD and the D-box both functionally and structurally, showing that a homodimer of ExbD captures one copy of the D-box peptide via beta-strand recruitment. We additionally show that both the D-box motif and ExbD are conserved in a range of Gram-negative bacteria, including members of the ESKAPE group of pathogens. The ExbD:D-box interaction is likely to represent an important aspect of force transduction between the inner and outer membranes. Given that TonB-dependent transport is an important contributor to virulence, this interaction is an intriguing potential target for novel antibacterial therapies.

A unique bacterial secretion machinery with multiple secretion centers.

The Porphyromonas gingivalis type IX secretion system (T9SS) promotes periodontal disease by secreting gingipains and other virulence factors. By in situ cryoelectron tomography, we report that the P. gingivalis T9SS consists of 18 PorM dimers arranged as a large, caged ring in the periplasm. Near the outer membrane, PorM dimers interact with a PorKN ring complex of ∼52 nm in diameter. PorMKN translocation complexes of a given T9SS adopt distinct conformations energized by the proton motive force, suggestive of different activation states. At the inner membrane, PorM associates with a cytoplasmic complex that exhibits 12-fold symmetry and requires both PorM and PorL for assembly. Activated motors deliver substrates across the outer membrane via one of eight Sov translocons arranged in a ring. The T9SSs are unique among known secretion systems in bacteria and eukaryotes in their assembly as supramolecular machines composed of apparently independently functioning translocation motors and export pores.

Direct interaction between fd phage pilot protein pIII and the TolQ-TolR proton-dependent motor provides new insights into the import of filamentous phages.

Filamentous phages are one of the simplest examples of viruses with a protein capsid that protects a circular single-stranded DNA genome. The infection is very specific, nonlytic, and can strongly affect the physiology or provide new pathogenic factors to its bacterial host. The infection process is proposed to rely on a pore-forming mechanism similar to that of certain nonenveloped eukaryotic viruses. The Ff coliphages (including M13, fd, and f1) have been intensively studied and were used to establish the sequence of events taking place for efficient crossing of the host envelope structure. However, the mechanism involved in the penetration of the cell inner membrane is not well understood. Here, we identify new host players involved in the phage translocation mechanism. Interaction studies by a combination of in vivo biochemical methods demonstrate that the adhesion protein pIII located at the tip of the phage binds to TolQ and TolR, two proteins that form a conserved proton-dependent molecular motor in the inner membrane of the host cell. Moreover, in vivo cysteine cross-linking studies reveal that the interactions between the pIII and TolQ or TolR occur between their transmembrane helix domains and may be responding to the proton motive force status of the cell. These results allow us to propose a model for the late stage of filamentous phage translocation mediated by multiple interactions with each individual component of the host TolQRA complex.

Molecular Genetic Dissection of the Regulatory Network of Proton Motive Force in Chloroplasts.

The proton motive force (pmf) generated across the thylakoid membrane rotates the Fo-ring of ATP synthase in chloroplasts. The pmf comprises two components: membrane potential (∆Ψ) and proton concentration gradient (∆pH). Acidification of the thylakoid lumen resulting from ∆pH downregulates electron transport in the cytochrome b6f complex. This process, known as photosynthetic control, is crucial for protecting photosystem I (PSI) from photodamage in response to fluctuating light. To optimize the balance between efficient photosynthesis and photoprotection, it is necessary to regulate pmf. Cyclic electron transport around PSI and pseudo-cyclic electron transport involving flavodiiron proteins contribute to the modulation of pmf magnitude. By manipulating the ratio between the two components of pmf, it is possible to modify the extent of photosynthetic control without affecting the pmf size. This adjustment can be achieved by regulating the movement of ions (such as K+ and Cl-) across the thylakoid membrane. Since ATP synthase is the primary consumer of pmf in chloroplasts, its activity must be precisely regulated to accommodate other mechanisms involved in pmf optimization. Although fragments of information about each regulatory process have been accumulated, a comprehensive understanding of their interactions is lacking. Here, I summarize current knowledge of the network for pmf regulation, mainly based on genetic studies.

Quantification of whole-organ individual and bilateral renal metabolic rate of oxygen.

PURPOSE: Renal metabolic rate of oxygen (rMRO2 ) is a potentially important biomarker of kidney function. The key parameters for rMRO2 quantification include blood flow rate (BFR) and venous oxygen saturation (SvO2 ) in a draining vessel. Previous approaches to quantify renal metabolism have focused on the single organ. Here, both kidneys are considered as one unit to quantify bilateral rMRO2 . A pulse sequence to facilitate bilateral rMRO2 quantification is introduced. METHODS: To quantify bilateral rMRO2 , measurements of BFR and SvO2 are made along the inferior vena cava (IVC) at suprarenal and infrarenal locations. From the continuity equation, these four parameters can be related to derive an expression for bilateral rMRO2 . The recently reported K-MOTIVE pulse sequence was implemented at four locations: left kidney, right kidney, suprarenal IVC, and infrarenal IVC. A dual-band variant of K-MOTIVE (db-K-MOTIVE) was developed by incorporating simultaneous-multi-slice imaging principles. The sequence simultaneously measures BFR and SvO2 at suprarenal and infrarenal locations in a single pass of 21 s, yielding bilateral rMRO2 . RESULTS: SvO2 and BFR are higher in suprarenal versus infrarenal IVC, and the renal veins are highly oxygenated (SvO2 >90%). Bilateral rMRO2 quantified in 10 healthy subjects (8 M, 30 ± 8 y) was found to be 291 ± 247 and 349 ± 300 (µmolO2 /min)/100 g, derived from K-MOTIVE and db-K-MOTIVE, respectively. In comparison, total rMRO2 from combining left and right was 329 ± 273 (µmolO2 /min)/100 g. CONCLUSION: The present work demonstrates that bilateral rMRO2 quantification is feasible with fair reproducibility and physiological plausibility. The indirect method is a promising approach to compute bilateral rMRO2 when individual rMRO2 quantification is difficult.

Fitness trade-offs of multidrug efflux pumps in Escherichia coli K-12 in acid or base, and with aromatic phytochemicals.

Multidrug efflux pumps are the frontline defense mechanisms of Gram-negative bacteria, yet little is known of their relative fitness trade-offs under gut conditions such as low pH and the presence of antimicrobial food molecules. Low pH contributes to the proton-motive force (PMF) that drives most efflux pumps. We show how the PMF-dependent pumps AcrAB-TolC, MdtEF-TolC, and EmrAB-TolC undergo selection at low pH and in the presence of membrane-permeant phytochemicals. Competition assays were performed by flow cytometry of co-cultured Escherichia coli K-12 strains possessing or lacking a given pump complex. All three pumps showed negative selection under conditions that deplete PMF (pH 5.5 with carbonyl cyanide 3-chlorophenylhydrazone or at pH 8.0). At pH 5.5, selection against AcrAB-TolC was increased by aromatic acids, alcohols, and related phytochemicals such as methyl salicylate. The degree of fitness cost for AcrA was correlated with the phytochemical's lipophilicity (logP). Methyl salicylate and salicylamide selected strongly against AcrA, without genetic induction of drug resistance regulons. MdtEF-TolC and EmrAB-TolC each had a fitness cost at pH 5.5, but salicylate or benzoate made the fitness contribution positive. Pump fitness effects were not explained by gene expression (measured by digital PCR). Between pH 5.5 and 8.0, acrA and emrA were upregulated in the log phase, whereas mdtE expression was upregulated in the transition-to-stationary phase and at pH 5.5 in the log phase. Methyl salicylate did not affect pump gene expression. Our results suggest that lipophilic non-acidic molecules select against a major efflux pump without inducing antibiotic resistance regulons.IMPORTANCEFor drugs that are administered orally, we need to understand how ingested phytochemicals modulate drug resistance in our gut microbiome. Bacteria maintain low-level resistance by proton-motive force (PMF)-driven pumps that efflux many different antibiotics and cell waste products. These pumps play a key role in bacterial defense by conferring resistance to antimicrobial agents at first exposure while providing time for a pathogen to evolve resistance to higher levels of the antibiotic exposed. Nevertheless, efflux pumps confer energetic costs due to gene expression and pump energy expense. The bacterial PMF includes the transmembrane pH difference (ΔpH), which may be depleted by permeant acids and membrane disruptors. Understanding the fitness costs of efflux pumps may enable us to develop resistance breakers, that is, molecules that work together with antibiotics to potentiate their effect. Non-acidic aromatic molecules have the advantage that they avoid the Mar-dependent induction of regulons conferring other forms of drug resistance. We show that different pumps have distinct selection criteria, and we identified non-acidic aromatic molecules as promising candidates for drug resistance breakers.

MRI-based quantification of whole-organ renal metabolic rate of oxygen.

During the early stages of diabetes, kidney oxygen utilization increases. The mismatch between oxygen demand and supply contributes to tissue hypoxia, a key driver of chronic kidney disease. Thus, whole-organ renal metabolic rate of oxygen (rMRO2 ) is a potentially valuable biomarker of kidney function. The key parameters required to determine rMRO2 include the renal blood flow rate (RBF) in the feeding artery and oxygen saturation in the draining renal vein (SvO2 ). However, there is currently no noninvasive method to quantify rMRO2 in absolute physiologic units. Here, a new MRI pulse sequence, Kidney Metabolism of Oxygen via T2 and Interleaved Velocity Encoding (K-MOTIVE), is described, along with evaluation of its performance in the human kidney in vivo. K-MOTIVE interleaves a phase-contrast module before a background-suppressed T2 -prepared balanced steady-state-free-precession (bSSFP) readout to measure RBF and SvO2 in a single breath-hold period of 22 s, yielding rMRO2 via Fick's principle. Variants of K-MOTIVE to evaluate alternative bSSFP readout strategies were studied. Kidney mass was manually determined from multislice gradient recalled echo images. Healthy subjects were recruited to quantify rMRO2 of the left kidney at 3-T field strength (N = 15). Assessments of repeat reproducibility and comparisons with individual measurements of RBF and SvO2 were performed, and the method's sensitivity was evaluated with a high-protein meal challenge (N = 8). K-MOTIVE yielded the following metabolic parameters: T2  = 157 ± 19 ms; SvO2  = 92% ± 6%; RBF = 400 ± 110 mL/min; and rMRO2  = 114 ± 117(µmol O2 /min)/100 g tissue. Reproducibility studies of T2 and RBF (parameters directly measured by K-MOTIVE) resulted in coefficients of variation less than 10% and intraclass correlation coefficients more than 0.75. The high-protein meal elicited an increase in rMRO2 , which was corroborated by serum biomarkers. The K-MOTIVE sequence measures SvO2 and RBF, the parameters necessary to quantify whole-organ rMRO2 , in a single breath-hold. The present work demonstrates that rMRO2 quantification is feasible with good reproducibility. rMRO2 is a potentially valuable physiological biomarker.

Proton conductance and regulation of proton/potassium fluxes in Escherichia coli FhlA-lacking cells during fermentation of mixed carbon sources.

Escherichia coli uptake potassium ions with the coupling of proton efflux and energy utilization via proton FOF1-ATPase. In this study contribution of formate hydrogen lyase (FHL) complexes in the proton/potassium fluxes and the formation of proton conductance (CMH+) were investigated using fhlA mutant strain. The proton flux rate (JH+) decreased in fhlA by âˆ¼ 25 % and ∼70 % during the utilization of glucose and glycerol, respectively, at 20 h suggesting H+ transport via or through FHL complexes. The decrease in JK+ in fhlA by ∼40 % proposed the interaction between FHL and Trk secondary transport system during mixed carbon fermentation. Moreover, the usage of N,N'-dicyclohexylcarbodiimide (DCCD) demonstrated the mediation of FOF1-ATPase in this interaction. CMH+ was 13.4 nmol min-1 mV-1 in WT at 20 h, which decreased by 20 % in fhlA. Taken together, FHL complexes have a significant contribution to the modulation of H+/K+ fluxes and the CMH + for efficient energy transduction and regulation of the proton motive force during mixed carbon sources fermentation.

Love Matters: The Effect of Mating Motive on Female Food Choice.

The prevalence of obesity has been increasing globally in recent decades. Behind the phenomenon, high-fat food consumption has been conceived as an important driver. In the current study, we explored whether mating motive caused an effect on female food choice as well as the psychological mechanism underlying it. In Study 1, we recruited 64 participants from a university and asked them to complete a mating prime, after which they would finish a food choice task in which food with different flavors were shown. In Study 2, we replicated Study 1 with a different mating priming method and examined the mediating role of body shaping desire on the relation between mating motive and female food choice. Results showed that: (1) The salience of mating motive decreased female's high-fat food choice but increased male's high-fat food choice; (2) the effect of mating motive in females was robust and more salient for sweet food rather than salty food; and (3) the body shaping desire partially mediated the effect of mating motive on female food choice.

Enhancing Escherichia coli abiotic stress resistance through ornithine lipid formation.

Escherichia coli is a common host for biotechnology and synthetic biology applications. During growth and fermentation, the microbes are often exposed to stress conditions, such as variations in pH or solvent concentrations. Bacterial membranes play a key role in response to abiotic stresses. Ornithine lipids (OLs) are a group of membrane lipids whose presence and synthesis have been related to stress resistance in bacteria. We wondered if this stress resistance could be transferred to bacteria not encoding the capacity to form OLs in their genome, such as E. coli. In this study, we engineered different E. coli strains to produce unmodified OLs and hydroxylated OLs by expressing the synthetic operon olsFC. Our results showed that OL formation improved pH resistance and increased biomass under phosphate limitation. Transcriptome analysis revealed that OL-forming strains differentially expressed stress- and membrane-related genes. OL-producing strains also showed better growth in the presence of the ionophore carbonyl cyanide 3-chlorophenylhydrazone (CCCP), suggesting reduced proton leakiness in OL-producing strains. Furthermore, our engineered strains showed improved heterologous violacein production at phosphate limitation and also at low pH. Overall, this study demonstrates the potential of engineering the E. coli membrane composition for constructing robust hosts with an increased abiotic stress resistance for biotechnology and synthetic biology applications. KEY POINTS: • Ornithine lipid production in E. coli increases biomass yield under phosphate limitation. • Engineered strains show an enhanced production phenotype under low pH stress. • Transcriptome analysis and CCCP experiments revealed reduced proton leakage.

Drinking contexts, coping motive, simultaneous cannabis use, and high-intensity drinking among adults in the United States.

AIMS: High-intensity drinking (HID), extreme drinking considerably above the level of heavy episodic drinking (HED), is associated with long-term health and social consequences. There is limited understanding of HID beyond young adulthood. This study aims to identify concurrent risk factors for HID, comparing age differences among all adults. METHODS: Multinomial logistic and linear regression modeling was performed using a nationally-representative sample of adults (analytic n = 7956) from the 2015 and 2020 National Alcohol Surveys. The outcomes were any HID of 8-11 drinks and 12+ drinks for men, and 8+ drinks for women, and corresponding frequencies. Concurrent risk factors included coping motive, sensation seeking, simultaneous use of alcohol and cannabis (SAC), and drinking at a bar or party. Analyses were stratified by age (18-29 vs. older) and sex. RESULTS: For younger men, sensation-seeking was significantly associated with HID (vs. no HED) at both levels and frequency of HID 8-11 drinks, while drinking to cope was only significant for 12+ drinks. For older men, drinking to cope was a consistent predictor for both HID level and its frequency, but sensation-seeking was not significant. Both coping and sensation-seeking were significantly associated with any HID for all women, while coping was significant for HID frequency for younger women. Frequent drinking at bars and parties were associated with greater odds of HID for all adults. With HED as referent, similar patterns of (though fewer significant) associations were observed. CONCLUSIONS: Younger and older adults share similar risk factors for HID, with coping more consistent for older men.

Different self-damaging behaviours, similar motives? Testing measurement invariance of motives for nonsuicidal self-injury, disordered eating and substance misuse.

OBJECTIVES: Theory and research suggest that distinct self-damaging behaviours (SDBs; e.g., nonsuicidal self-injury [NSSI], restrictive eating, binge eating, drug misuse, alcohol misuse) share similar motives. However, few studies have used a common self-report inventory to investigate the shared relevance and relative salience of motives for SDBs. Accordingly, the present study: (1) examined whether self-report scales assessing intrapersonal motives (i.e., relieving negative emotions, enhancing positive emotions, punishing oneself) and interpersonal motives (i.e., bonding with others, conforming with others, communicating distress, communicating strength, reducing demands) have invariant factor structures across SDBs; and (2) compared the salience of these motives across SDBs. METHODS: 1018 adults (54.6% men, Mage = 35.41 years) with a history of SDBs were allocated to the following groups: NSSI (n = 213), restrictive eating (n = 200), binge eating (n = 200), drug misuse (n = 200) or alcohol misuse (n = 205). Participants reported on their motives for engaging in their allocated SDB. Measurement invariance analyses compared the factor structures and latent means of the motive scales across SDBs. RESULTS: The motive scales had comparable factor structures across SDBs. Intrapersonal motives were most strongly endorsed for NSSI and drug misuse. Interpersonal motives were most strongly endorsed for drug and alcohol misuse. All motives were least salient to restrictive eating. CONCLUSIONS: Results suggest that common motives underlie distinct SDBs and that they can be adequately assessed using a single self-report inventory. However, certain motives are more relevant to some SDBs than others, with restrictive eating being the most motivationally distinct SDB. This knowledge can inform transdiagnostic models and interventions for SDBs.

Diet quality and associations with motivation and ability to consume a healthy diet among adolescents from urban low-income households in Bangladesh.

In low- and middle-income countries, particularly in urban areas, adolescent diets consist mainly of energy-dense and nutrient-poor foods, putting them at risk of malnutrition and non-communicable diseases (NCD). In Bangladesh, little is known about the diet quality of adolescents, their food choices and the drivers of such choices. This study assessed motivations and ability to consume a healthy diet among adolescent girls and boys from low-income urban families and how these drivers were associated with dietary diversity and diet quality. A cross-sectional survey was conducted among 299 adolescents (15-19 years) from low-income households in Dhaka city during September-October 2020. The Diet Quality Questionnaire was used to collect non-quantitative food intake in the previous day or night to calculate diet quality indicators of food group diversity score, % of adolescents achieving minimum dietary diversity, NCD-Protect and NCD-Risk and the Global Dietary Recommendations score. Motivation was measured by 11 food choice motives. Ability was measured by belief in own ability to engage in healthy eating behaviors (self-efficacy). Adolescent diets showed a mean food group diversity of 4.9 out of 10, with 60% of adolescents achieving minimum dietary diversity, but lacked health-promoting foods (average of 2.7 out of 9 food groups) yet included few foods to avoid and limit (1.6 out of 9). Adolescents valued food choice motive 'safety' the most, followed by 'health', 'taste', 'price', 'convenience' and 'local or seasonal'. A higher motivation to consume 'local or seasonal' and a lower motivation driven by 'price', and a higher perceived self-efficacy were associated with better diet quality. Future interventions should address self-efficacy, concerns about food price and increase local and seasonal foods availability in the urban poor food environment of Dhaka to improve overall diet quality.

Membrane voltage-dependent activation mechanism of the bacterial flagellar protein export apparatus.

The proton motive force (PMF) consists of the electric potential difference (Δψ), which is measured as membrane voltage, and the proton concentration difference (ΔpH) across the cytoplasmic membrane. The flagellar protein export machinery is composed of a PMF-driven transmembrane export gate complex and a cytoplasmic ATPase ring complex consisting of FliH, FliI, and FliJ. ATP hydrolysis by the FliI ATPase activates the export gate complex to become an active protein transporter utilizing Δψ to drive proton-coupled protein export. An interaction between FliJ and a transmembrane ion channel protein, FlhA, is a critical step for Δψ-driven protein export. To clarify how Δψ is utilized for flagellar protein export, we analyzed the export properties of the export gate complex in the absence of FliH and FliI. The protein transport activity of the export gate complex was very low at external pH 7.0 but increased significantly with an increase in Δψ by an upward shift of external pH from 7.0 to 8.5. This observation suggests that the export gate complex is equipped with a voltage-gated mechanism. An increase in the cytoplasmic level of FliJ and a gain-of-function mutation in FlhA significantly reduced the Δψ dependency of flagellar protein export by the export gate complex. However, deletion of FliJ decreased Δψ-dependent protein export significantly. We propose that Δψ is required for efficient interaction between FliJ and FlhA to open the FlhA ion channel to conduct protons to drive flagellar protein export in a Δψ-dependent manner.

Nursing students' approaches to learning in selected Malawian nursing schools: a cross-sectional study.

BACKGROUND: Students' approaches to learning are of essence in nursing education. This is because nursing is a profession where classroom learning leads to clinical performance. Although the literature recognizes student's approaches to learning as a significant aspect affecting the quality of students' learning, studies suggest that quality of learning has not been highly achieved in Malawian nursing colleges. Currently, there is a scarcity of empirical data on the learning approaches that Malawian nursing and midwifery students in nursing colleges employ. This study assessed the different approaches to learning among nursing and midwifery students in selected Malawian nursing colleges. METHODS: This was a cross- sectional study that employed quantitative methods. The target population was nursing and midwifery students pursuing nursing diplomas from Nkhoma College of Nursing, Ekwendeni College of Health Sciences and Malawi College of Health Sciences. A total of 251 students were sampled randomly from the three nursing colleges. Data was collected through a self-administered questionnaire (R-SPQ-2 F) by Biggs. The data was analyzed using chi-square and binary logistic regression. In this study Cronbach's alpha was 0.6. RESULTS: Most students had used a deep approach to learning (M = 3.201, SD = 0.623) than the surface approach (M = 2.757, SD = 0.732). Being in the age category of 16-20 had more likelihood of adopting a surface approach to learning compared to other age categories (X2 = 7.669, DF 2, P = .02). Students from Malawi College of Health Sciences were more likely to adopt a surface approach to learning compared to students from Nkhoma Nursing College and Ekwendeni College of Health Sciences (X2 = 12.388, df = 2, P = .002). CONCLUSION: A deep approach to learning emerged as the most preferred approach to learning which indirectly implies that most students attain meaningful learning. Age and environment are some of the key determinants associated with different learning approaches. More attention should be given to younger students during teaching and learning to promote deep learning.

Fermentation Revisited: How Do Microorganisms Survive Under Energy-Limited Conditions?

During fermentation FOF1 hydrolyzes ATP, coupling proton transport to proton-motive force (pmf) generation. Despite that, pmf generated by ATP hydrolysis does not satisfy the energy budget of a fermenting cell. However, pmf can also be generated by extrusion of weak organic acids such as lactate and by hydrogen cycling catalyzed by hydrogenases (Hyds). Here we highlight recent advances in our understanding of how the transport of weak organic acids and enzymes contributes to pmf generation during fermentation. The potential impact of these processes on metabolism and energy conservation during microbial fermentation have been overlooked and they not only expand on Mitchell's chemiosmotic theory but also are of relevance to the fields of microbial biochemistry and human and animal health.