Structure of detergent-activated BAK dimers derived from the inert monomer.

A body of data supports the existence of core (α2-α5) dimers of BAK and BAX in the oligomeric, membrane-perturbing conformation of these essential apoptotic effector molecules. Molecular structures for these dimers have only been captured for truncated constructs encompassing the core domain alone. Here, we report a crystal structure of BAK α2-α8 dimers (i.e., minus its flexible N-terminal helix and membrane-anchoring C-terminal segment) that has been obtained through the activation of monomeric BAK with the detergent C12E8. Core dimers are evident, linked through the crystal by contacts via latch (α6-α8) domains. This crystal structure shows activated BAK dimers with the extended latch domain present. Our data provide direct evidence for the conformational change converting BAK from inert monomer to the functional dimer that destroys mitochondrial integrity. This dimer is the smallest functional unit for recombinant BAK or BAX described so far.

Post-infection treatment with the E protein inhibitor BIT225 reduces disease severity and increases survival of K18-hACE2 transgenic mice infected with a lethal dose of SARS-CoV-2.

The Coronavirus envelope (E) protein is a small structural protein with ion channel activity that plays an important role in virus assembly, budding, immunopathogenesis and disease severity. The viroporin E is also located in Golgi and ER membranes of infected cells and is associated with inflammasome activation and immune dysregulation. Here we evaluated in vitro antiviral activity, mechanism of action and in vivo efficacy of BIT225 for the treatment of SARS-CoV-2 infection. BIT225 showed broad-spectrum direct-acting antiviral activity against SARS-CoV-2 in Calu3 and Vero cells with similar potency across 6 different virus strains. BIT225 inhibited ion channel activity of E protein but did not inhibit endogenous currents or calcium-induced ion channel activity of TMEM16A in Xenopus oocytes. BIT225 administered by oral gavage for 12 days starting 12 hours before infection completely prevented body weight loss and mortality in SARS-CoV-2 infected K18 mice (100% survival, n = 12), while all vehicle-dosed animals reached a mortality endpoint by Day 9 across two studies (n = 12). When treatment started at 24 hours after infection, body weight loss, and mortality were also prevented (100% survival, n = 5), while 4 of 5 mice maintained and increased body weight and survived when treatment started 48 hours after infection. Treatment efficacy was dependent on BIT225 dose and was associated with significant reductions in lung viral load (3.5 log10), virus titer (4000 pfu/ml) and lung and serum cytokine levels. These results validate viroporin E as a viable antiviral target and support the clinical study of BIT225 for treatment and prophylaxis of SARS-CoV-2 infection.

The energetics and evolution of oxidoreductases in deep time.

The core metabolic reactions of life drive electrons through a class of redox protein enzymes, the oxidoreductases. The energetics of electron flow is determined by the redox potentials of organic and inorganic cofactors as tuned by the protein environment. Understanding how protein structure affects oxidation-reduction energetics is crucial for studying metabolism, creating bioelectronic systems, and tracing the history of biological energy utilization on Earth. We constructed ProtReDox (https://protein-redox-potential.web.app), a manually curated database of experimentally determined redox potentials. With over 500 measurements, we can begin to identify how proteins modulate oxidation-reduction energetics across the tree of life. By mapping redox potentials onto networks of oxidoreductase fold evolution, we can infer the evolution of electron transfer energetics over deep time. ProtReDox is designed to include user-contributed submissions with the intention of making it a valuable resource for researchers in this field.

Galectin-3 N-terminal tail prolines modulate cell activity and glycan-mediated oligomerization/phase separation.

Galectin-3 (Gal-3) has a long, aperiodic, and dynamic proline-rich N-terminal tail (NT). The functional role of the NT with its numerous prolines has remained enigmatic since its discovery. To provide some resolution to this puzzle, we individually mutated all 14 NT prolines over the first 68 residues and assessed their effects on various Gal-3-mediated functions. Our findings show that mutation of any single proline (especially P37A, P55A, P60A, P64A/H, and P67A) dramatically and differentially inhibits Gal-3-mediated cellular activities (i.e., cell migration, activation, endocytosis, and hemagglutination). For mechanistic insight, we investigated the role of prolines in mediating Gal-3 oligomerization, a fundamental process required for these cell activities. We showed that Gal-3 oligomerization triggered by binding to glycoproteins is a dynamic process analogous to liquid-liquid phase separation (LLPS). The composition of these heterooligomers is dependent on the concentration of Gal-3 as well as on the concentration and type of glycoprotein. LLPS-like Gal-3 oligomerization/condensation was also observed on the plasma membrane and disrupted endomembranes. Molecular- and cell-based assays indicate that glycan binding-triggered Gal-3 LLPS (or LLPS-like) is driven mainly by dynamic intermolecular interactions between the Gal-3 NT and the carbohydrate recognition domain (CRD) F-face, although NT-NT interactions appear to contribute to a lesser extent. Mutation of each proline within the NT differentially controls NT-CRD interactions, consequently affecting glycan binding, LLPS, and cellular activities. Our results unveil the role of proline polymorphisms (e.g., at P64) associated with many diseases and suggest that the function of glycosylated cell surface receptors is dynamically regulated by Gal-3.

Experiential avoidance as a mediator of risk factors for higher order internalizing psychopathology in the perinatal period.

OBJECTIVES: Perinatal psychopathology can be damaging. This study examined the strength of the associations between risk factors and all perinatal mood and anxiety disorder symptoms while assessing the mediating effect of experiential avoidance. METHOD: Participants (N = 246) completed assessments during pregnancy (28-32 weeks) and the postpartum (6-8 weeks). Structural equation modeling (SEM) was used to examine associations between risk factors and latent factors: distress (composed of depression, generalized anxiety, irritability, and panic symptoms); fear (social anxiety, agoraphobia, specific phobia, and obsessive-compulsive); and bipolar (mania and obsessive-compulsive). RESULTS: During pregnancy, past psychiatric history, anxiety sensitivity, maladaptive coping, and age were significant risk factors. In the postpartum, negative maternal attitudes and past psychiatric history were only risk factors for symptoms that composed distress. Experiential avoidance mediated the relation between maladaptive coping and symptoms that composed fear. CONCLUSION: It is important to assess for psychological risk factors starting in pregnancy. This study identified critical risk factors that are associated with the underlying commonality among perinatal mood and anxiety symptoms. Some of the risk factors as well as the mediator are malleable (negative maternal attitudes, experiential avoidance), creating new possibilities for prevention and treatment of perinatal mood and anxiety disorder symptoms.

Validation of Anti-Adeno Associated Virus Serotype rh10 (AAVrh.10) Total and Neutralizing Antibody Immunogenicity Assays.

Immunogenicity assessment of Adeno-Associated Virus (AAV) vectors is a critical part of gene therapy drug development. Whether the assays are used for inclusion/exclusion criteria or to monitor the safety and efficacy of the gene therapy, they are critical bioanalytical assessments. While total anti-AAV assays are perceived as easier to develop and implement than neutralizing anti-AAV assays, the gene therapy field is still nascent, and it is not yet clear which of the assays should be implemented at what stage of drug development. Recently AAVrh.10 has gained interest for use in gene therapies targeting cardiac, neurological, and other diseases due to its enhanced transduction efficiency. There is limited information on anti-AAVrh.10 antibodies and their clinical impact; thus, the information presented herein documents the validation of both a total antibody assay (TAb) and a neutralizing antibody (NAb) assay for anti-AAVrh.10 antibodies. In this manuscript, the validation was performed in accordance with the 2019 FDA immunogenicity guidance with additional evaluations to comply with CLIA where applicable. The AAVrh.10 TAb and NAb assays were compared in terms of sensitivity, drug tolerance, and precision, along with a concordance analysis using the same individual serum samples. This comparison gave insight into the utility of each format as a screening assay for inclusion into clinical studies.

The ability of the Global Leadership Initiative on Malnutrition (GLIM) to diagnose protein-energy malnutrition in patients requiring vascular surgery: a validation study.

Identifying nutritional deficits and implementing appropriate interventions in patients requiring vascular surgery is challenging due to the paucity of appropriate screening and assessment tools in this group. This retrospective study aimed to determine the validity of the Global Leadership Initiative on Malnutrition (GLIM) in identifying protein-energy malnutrition (PEM) in inpatients admitted to a vascular surgery unit, using the PG-SGA as the comparator. Diagnostic accuracy and consistency were determined between the GLIM and the Patient-Generated Subjective Global Assessment (PG-SGA) global rating. The GLIM determination was made retrospectively using the relevant parameters collected at baseline in the original study. Two hundred and twenty-four (70·1 % male) participants were included. The prevalence of PEM was 28·6 % on GLIM and 17 % via the PG-SGA. Compared with the PG-SGA, the GLIM achieved sensitivity of 73·7 % and specificity of 80·6 %; however positive predictive value was 43·7 % indicating that the GLIM over-diagnosed malnutrition compared with the PG-SGA. Kappa reached 0·427 indicating moderate diagnostic consistency. Due to the absence of an ideal instrument and the complexity of malnutrition often seen in this group which extends beyond PEM to significant micronutrient deficiencies, further work is required to determine the most appropriate instrument in this patient group, and how micronutrient status can also be included in the overall assessment given the critical role of micronutrients in this group.

Postpartum care needs assessment: women's understanding of postpartum care, practices, barriers, and educational needs.

BACKGROUND: Complications in the postpartum period pose substantial risks to women and can result in significant maternal morbidity and mortality. However, there is much less attention on postpartum care compared to pregnancy and childbirth. The goal of this study was to gather information on women's knowledge of postpartum care and complications, recovery practices after childbirth, perceived barriers to receiving care during the postpartum period, and educational needs in four health centers. The findings can inform the development of appropriate curriculum and interventions for postnatal care education in similar settings. METHODS: A descriptive qualitative study design was employed. Eight focus group discussions were conducted among 54 postpartum women who delivered in four health centers in Sagnarigu District in Tamale, Ghana. Audio recordings of focus group data were transcribed and translated, and thematic analysis was conducted. RESULTS: There were six main themes that emerged from the focus group discussions: 1) baby focused postpartum care; 2) postpartum practices; 3) inadequate knowledge ofpostpartum danger signs; 4) barriers to accessing postpartum care 5) experiences of poor mental health; and 6) need for postpartum education. CONCLUSIONS: Postpartum care for women in this study was primarily perceived as care of the baby post-delivery and missing key information on physical and mental health care for the mother. This can result in poor adjustment postpartum and critically, a lack of knowledge on danger signs for common causes of morbidity and mortality in the postpartum period. Future research needs to understand how to communicate important information on postpartum mental and physical health to better protect mothers in the region.

An exploratory study into the relationship between playing at home or away and concussion.

PRIMARY OBJECTIVE: To investigate the effect of home and away game travel on risk of concussion across different levels of rugby union. RESEARCH DESIGN: Exploration study across school, university, and professional rugby teams. METHODS AND PROCEDURES: Retrospective analysis of concussion incidence and symptomology of surveillance data and prospective data collection for potential concussions via surveys. Data was collected from school rugby teams (n = 344 matches, over 2 years), a university rugby (n = 6 matches), and a professional rugby team (n = 64 matches, over two seasons). MAIN OUTCOMES AND RESULTS: School level rugby had an increased prevalence of concussions in away matches (p = 0.02). Likewise, there was a significant increase (p < 0.05) in concussions at away matches in university rugby. In addition, the professional rug by team had significant differences in recovery times and symptoms with away fixtures, including longer recovery times (p < 0.01), more initial symptoms (p < 0.01), as well as greater and more severe symptoms at 48 hours (p < 0.05). CONCLUSIONS: This research highlights an increased prevalence of concussion in school and university-aged rugby players away from home, as well as increased symptoms, symptom severity, and recovery times in professional rugby players.

Low muscle mass determined by psoas muscle area does not correlate with dual-energy x-ray absorptiometry or total lumbar muscle mass scores: A prospective cohort study of patients undergoing vascular surgery.

Background/Objective: Low muscle mass and sarcopenia have been explored as risk factors for poor outcomes following vascular surgery. The findings have been variable. The use of a diverse range of techniques to identify low muscle mass is a confounder in establishing the true relationship between low muscle mass, sarcopenia and outcomes. Our aim was to establish if different scoring methods identified the same patients as sarcopenic. We also explored which method best predicted outcomes. Method: 70 patients undergoing vascular surgery were prospectively assessed for sarcopenia using dual-energy x-ray absorptiometry (DEXA) scan, grip strength and gait speed. These patients underwent abdominal CT imaging as routine care. The muscle mass of each patient was determined using DEXA and by both psoas muscle and total skeletal muscle area on CT, normalised for patient height (PMI and CT-SMI, respectively). Low muscle mass was defined by published age- and sex-specific cut-offs. Grip strength data was combined with muscle mass to define sarcopenic patients. One- and 3-year mortality and time to readmission was recorded. Conclusion and Results: 10-22% of patients had low muscle mass and 4-10% of patients were sarcopenic, depending on the method employed. PMI did not correlate with DEXA or CT-SMI for low muscle mass, but CT-SMI correlated with DEXA (p = 0.0007). For sarcopenia, CT-SMI and DEXA scoring correlated (p = 0.002); PMI correlated with CT-SMI (p = 0.0006) but not DEXA. Low muscle mass by PMI predicted 1-year mortality (p = 0.02, X2 = 5.34, Effect size = 1.04) but the applicability of this finding is limited by the diverse pathologies explored. No other method predicted 1- or 3-year mortality or readmissions in this heterogenous cohort. The psoas area did not correlate with muscle mass defined by DEXA or total lumbar skeletal muscle area. Low psoas muscle index may be an independent marker of poor outcome, unrelated to generalised sarcopenia, and this warrants investigation in specific pathologies. A lower total number of patients were sarcopenic than had been expected, emphasising the need to use population-based pre-defined cut-offs.

Chemokines and galectins form heterodimers to modulate inflammation.

Chemokines and galectins are simultaneously upregulated and mediate leukocyte recruitment during inflammation. Until now, these effector molecules have been considered to function independently. Here, we tested the hypothesis that they form molecular hybrids. By systematically screening chemokines for their ability to bind galectin-1 and galectin-3, we identified several interacting pairs, such as CXCL12 and galectin-3. Based on NMR and MD studies of the CXCL12/galectin-3 heterodimer, we identified contact sites between CXCL12 ß-strand 1 and Gal-3 F-face residues. Mutagenesis of galectin-3 residues involved in heterodimer formation resulted in reduced binding to CXCL12, enabling testing of functional activity comparatively. Galectin-3, but not its mutants, inhibited CXCL12-induced chemotaxis of leukocytes and their recruitment into the mouse peritoneum. Moreover, galectin-3 attenuated CXCL12-stimulated signaling via its receptor CXCR4 in a ternary complex with the chemokine and receptor, consistent with our structural model. This first report of heterodimerization between chemokines and galectins reveals a new type of interaction between inflammatory mediators that can underlie a novel immunoregulatory mechanism in inflammation. Thus, further exploration of the chemokine/galectin interactome is warranted.

Obsessive-compulsive and related disorder symptoms in the perinatal period: prevalence and associations with postpartum functioning.

Obsessive-compulsive disorder (OCD) symptoms are more likely to develop or be exacerbated during pregnancy and the postpartum period, which can cause significant distress and impairment. However, the disorders grouped with OCD in the DSM-5, obsessive-compulsive and related disorders (OCRD; e.g., hoarding disorder (HD), body dysmorphic disorder (BDD), trichotillomania (TTM), excoriation disorder (ED)), have rarely been examined in the perinatal period. This study aimed to explore (1) the prevalence of all clinically significant OCRD symptoms in pregnancy and the postpartum period and (2) the correlations between OCRD psychopathology and postpartum functioning. Participants were recruited during their second trimester of pregnancy from a Midwestern medical center. Participants completed an online questionnaire and a semi-structured clinical interview during pregnancy (28-32 weeks' gestation, N = 276) and the postpartum period (6-8 weeks, N = 221). BDD and OCD symptoms were the most prevalent. In pregnancy, 14.9% (N = 41) of participants endorsed clinically significant BDD symptoms and 6.2% (N = 17) endorsed clinically significant OCD symptoms. In the postpartum period, 11.8% (N = 26) endorsed clinically significant BDD symptoms and 14% (N = 31) endorsed clinically significant OCD symptoms. Poorer postpartum functioning was associated with elevated OCRD symptoms in pregnancy and postpartum. OCRD symptoms occur during pregnancy and the postpartum period at rates similar or higher than other life periods. Elevated OCRD symptoms are associated with poorer postpartum functioning across domains. Future research should explore how all OCRD symptoms may affect functioning in the perinatal period, not only OCD symptoms.

Physical activity interventions in older people with cancer: A review of systematic reviews.

INTRODUCTION: Whilst there has been a wealth of research on benefits of physical activity (PA) in people with cancer, with three published reviews of reviews, no review of reviews has focused on older adults (65 years or older) who may have unique biological characteristics and barriers. We summarised PA effectiveness from reviews where majority of study participants were 65 years or older. METHODS: Six databases were searched for systematic reviews of randomised controlled studies (RCTs)/quasi-RCTs examining any type of PA in reviews where majority of study participants were aged 65 years or older. Two reviewers conducted the search and analysis according to PRISMA and JBI guidelines. RESULTS: Fifteen reviews involving 76 different primary studies (5404 participants) were included. The majority (3827; 71%) had prostate cancer. PA was associated with benefits across multiple physical outcomes (muscle mass, functional performance, strength), improved fatigue and health service outcomes. In contrast to younger adults, there was no improvement in anxiety and mixed findings for quality of life and depression. CONCLUSION: PA is associated with multiple benefits in older adults with cancer, with some differences compared to younger individuals which may reflect biological or behavioural determinants. Future research should focus on mechanisms underlying PA effectiveness and underrepresented populations.

Treating posttraumatic stress disorder at home in a single week using 1-week virtual massed cognitive processing therapy.

Posttraumatic stress disorder (PTSD) treatments are increasingly delivered in massed formats and have shown comparable results to standard, weekly treatment. To date, massed cognitive processing therapy (CPT), delivered daily, has been delivered primarily in combination with adjunctive services and among veteran populations, but it has not been rigorously evaluated as a standalone intervention. The present study evaluated 1-week massed CPT delivered virtually (i.e., via telehealth) to a community sample of trauma-exposed individuals (N = 24). Using a single-arm open-label design, participants received CPT twice per day for 5 days. The results indicated that most participants completed treatment (n = 23, 95.8%), and no adverse events were reported. Participants exhibited large reductions in clinician-rated, d = 2.01, and self-reported PTSD symptoms, d = 2.55, as well as self-reported depressive symptoms, d = 1.46. On average, participants reported a 5-point PTSD symptom reduction and 1-point reduction in depressive symptoms for each treatment day. Reductions in PTSD and depressive symptoms were maintained at 3-month follow-up. Overall, 1-week massed CPT delivered virtually was shown to be feasible and to result in rapid symptom reductions that were sustained over time. Virtual massed CPT has the potential to increase access to effective treatments and help trauma survivors restore aspects of their lives in short amounts of time.

Resilience of T cell-intrinsic dysfunction in transplantation tolerance.

Following antigen stimulation, naïve T cells differentiate into memory cells that mediate antigen clearance more efficiently upon repeat encounter. Donor-specific tolerance can be achieved in a subset of transplant recipients, but some of these grafts are rejected after years of stability, often following infections. Whether T cell memory can develop from a tolerant state and whether these formerly tolerant patients develop antidonor memory is not known. Using a mouse model of cardiac transplantation in which donor-specific tolerance is induced with costimulation blockade (CoB) plus donor-specific transfusion (DST), we have previously shown that systemic infection with Listeria monocytogenes (Lm) months after transplantation can erode or transiently abrogate established tolerance. In this study, we tracked donor-reactive T cells to investigate whether memory can be induced when alloreactive T cells are activated in the setting of tolerance. We show alloreactive T cells persist after induction of cardiac transplantation tolerance, but fail to acquire a memory phenotype despite becoming antigen experienced. Instead, donor-reactive T cells develop T cell-intrinsic dysfunction evidenced when removed from the tolerant environment. Notably, Lm infection after tolerance did not rescue alloreactive T cell memory differentiation or functionality. CoB and antigen persistence were sufficient together but not separately to achieve alloreactive T cell dysfunction, and conventional immunosuppression could substitute for CoB. Antigen persistence was required, as early but not late surgical allograft removal precluded the acquisition of T cell dysfunction. Our results demonstrate transplant tolerance-associated T cell-intrinsic dysfunction that is resistant to memory development even after Lm-mediated disruption of tolerance.

Climate change and Australia's primary industries: factors hampering an effective and coordinated response.

Australia's primary production sector operates in one of the world's most variable climates with future climate change posing a challenge to its ongoing sustainability. Recognising this, Australia has invested in understanding climate change risks to primary production with a substantial amount of research produced. Recently, focus on this research space has broadened, with interests from the financial sector and expanded scopes of works from government and industry. These expanded needs require sector- and country-wide assessments to assist with the implementation of climate strategies. We considered the applicability of the current research body for these needs by reviewing 188 peer-reviewed studies that considered the quantitative impacts of climate change on Australia's primary industries. Our broad review includes cropping, livestock, horticulture, forestry and fisheries and biosecurity threats. This is the first such review for Australia, and no other similar country-wide review was found. We reviewed the studies through three lenses, industry diversity, geographic coverage and study comparability. Our results show that all three areas are lacking for sector- and country-wide assessments. Industry diversity was skewed towards cropping and biosecurity threats (64% of all studies) with wheat in particular a major focus (25% of all studies). Geographic coverage at a state level appeared to be evenly distributed across the country; however, when considered in conjunction with industry focus, gaps emerged. Study comparability was found to be very limited due to the use of different historical baseline periods and different impact models. We make several recommendations to assist with future research directions, being (1) co-development of a standard set of method guidelines for impact assessments, (2) filling industry and geographic knowledge gaps, and (3) improving transparency in study method descriptions. Uptake of these recommendations will improve study application and transparency enabling and enhancing responses to climate change in Australia's primary industries.

Communal mastery and associations with depressive and PTSD symptomatology among urban trauma-exposed women.

Objective: Racial and ethnic minority women from low-resource urban communities experience disproportionately high rates of trauma exposure. Higher rates of lifetime trauma exposure are strongly associated with subsequent psychological sequela, specifically depression and posttraumatic stress disorder (PTSD). Communal mastery is the ability to cope with challenges and achieve goals by being closely interconnected with friends, family, and significant others. Yet, it is unknown if communal mastery is protective specifically against PTSD and depressive symptoms. Method: Participants (N = 131) were Black and Latina women (88.5% Black, mean monthly income: < $750) recruited from an urban outpatient obstetric-gynecological clinic at an academic medical center. Participants completed an online questionnaire that assessed trauma history, PTSD and depressive symptoms, types of individualistic coping, social support, and communal mastery. Results: Hierarchical multiple regression models demonstrated that communal mastery is uniquely associated with fewer PTSD symptoms (ß = -.23, p = .003). More severe trauma history, more use of passive coping skills, and poorer social support were also significantly associated with PTSD symptoms, explaining over half of the variance in PTSD symptoms. Although significantly correlated, communal mastery was not uniquely associated with fewer depressive symptoms (ß = -.13, p = .201). Conclusions: These findings suggest that connectedness as assessed through communal mastery serves as an important shield against the effects of traumatic stress for Black and Latina women. Future research would benefit by exploring interventions that aim to increase communal mastery in order to help highly trauma-exposed racial and ethnic minority women in low-resource environments. (PsycInfo Database Record (c) 2022 APA, all rights reserved).

The methyltransferase G9a regulates HoxA9-dependent transcription in AML.

Chromatin modulators are emerging as attractive drug targets, given their widespread implication in human cancers and susceptibility to pharmacological inhibition. Here we establish the histone methyltransferase G9a/EHMT2 as a selective regulator of fast proliferating myeloid progenitors with no discernible function in hematopoietic stem cells (HSCs). In mouse models of acute myeloid leukemia (AML), loss of G9a significantly delays disease progression and reduces leukemia stem cell (LSC) frequency. We connect this function of G9a to its methyltransferase activity and its interaction with the leukemogenic transcription factor HoxA9 and provide evidence that primary human AML cells are sensitive to G9A inhibition. Our results highlight a clinical potential of G9A inhibition as a means to counteract the proliferation and self-renewal of AML cells by attenuating HoxA9-dependent transcription.

PLG-007 and Its Active Component Galactomannan-α Competitively Inhibit Enzymes That Hydrolyze Glucose Polymers.

PLG-007 is a developmental therapeutic compound that has been clinically shown to reduce the magnitude of postprandial glucose excursions and has the potential to be an adjunct treatment for diabetes and inflammatory-related diseases. The present investigation is aimed at understanding the molecular mechanism of action of PLG-007 and its galactomannan (GM) components GMα and GMß (in a 1:4 mass ratio, respectively) on enzyme (i.e., α-amylase, maltase, and lactase) hydrolysis of glucose polymers using colorimetric assays and 13C HSQC NMR spectroscopy. The starch-iodine colorimetric assay indicated that GMα strongly inhibits α-amylase activity (~16-fold more potent than GMß) and thus is the primary active component in PLG-007. 13C HSQC experiments, used to follow the α-amylase-mediated hydrolysis of starch and amylopectin, further demonstrate the α-amylase inhibitory effect of GMα via α-amylase-mediated hydrolysis of starch and amylopectin. Maltohexaose (MT6) was used to circumvent the relative kinetic complexity of starch/amylopectin degradation in Michaelis-Menten analyses. The Vmax, KM, and Ki parameters were determined using peak volume integrals from 13C HSQC NMR spectra. In the presence of PLG-007 with α-amylase and MT6, the increase in KM from 7.5 ± 0.6 × 10-3 M (control) to 21 ± 1.4 × 10-3 M, with no significant change in Vmax, indicates that PLG-007 is a competitive inhibitor of α-amylase. Using KM values, Ki was estimated to be 2.1 ± 0.9 × 10-6 M; however, the microscopic Ki value of GMα is expected to be larger as the binding stoichiometry is likely to be greater than 1:1. Colorimetric assays also demonstrated that GMα is a competitive inhibitor of the enzymes maltase and lactase. Overall, this study provides insight as to how PLG-007 (GMα) is likely to function in vivo.

Selective border permeability: Governing complex environmental issues through and beyond COVID-19.

COVID-19 has changed the permeability of borders in transboundary environmental governance regimes. While borders have always been selectively permeable, the pandemic has reconfigured the nature of cross-border flows of people, natural resources, finances and technologies. This has altered the availability of spaces for enacting sustainability initiatives within and between countries. In Southeast Asia, national governments and businesses seeking to expedite economic recovery from the pandemic-induced recession have selectively re-opened borders by accelerating production and revitalizing agro-export growth. Widening regional inequities have also contributed to increased cross-border flows of illicit commodities, such as trafficked wildlife. At the same time, border restrictions under the exigencies of controlling the pandemic have led to a rolling back and scaling down of transboundary environmental agreements, regulations and programs, with important implications for environmental democracy, socio-ecological justice and sustainability. Drawing on evidence from Southeast Asia, the article assesses the policy challenges and opportunities posed by the shifting permeability of borders for organising and operationalising environmental activities at different scales of transboundary governance.