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This longitudinal quasi-experimental study examines the relationship between social class, subjective health, motivation to return to work (RTW) following medical rehabilitation, and participation in aftercare interventions, utilizing the Social Cognitive Theory of Social Class. To do so, a prospective multicentric study was conducted. The analysis was based on data from N = 1044 orthopedic rehabilitation patients in three clinics in Germany. Latent growth curve models, fixed-effect linear regression models, and multilevel binomial logistic regression were employed for data analysis. As hypothesized, the findings demonstrate that lower social class is associated with poorer subjective health, while higher social class is linked to increased solipsistic motives, characterized by pursuing personal goals in the context of RTW. Conversely, individuals from lower social classes exhibit contextualist motives, which indicate a focus on social and environmental threats influencing their motivation to RTW. Surprisingly, social class does not significantly impact participation in aftercare interventions, probably due to low variance and potentially the successful inclusion within the German healthcare system. These findings emphasize the importance of considering diverse motivation profiles derived from the Social Cognitive Theory of Social Class. The study contributes to our understanding of the social determinants of health and has implications for reducing health disparities by highlighting the motivational aspects, including solipsistic and contextualist motives, associated with social class. • Social class shapes subjective health trajectories in orthopedic rehabilitation patients. • Individuals from lower social classes initially exhibit lower subjective health. • Rehabilitation may equalize health improvements across social classes. • Solipsistic and contextualist return-to-work motives may vary based on social class. • No class differences in participation in aftercare programs were observed. [ABSTRACT FROM AUTHOR]

• People with preexisting anxiety-related disorders are vulnerable to a steep inflation of fear during COVID-19 pandemic. • Cognitive behavioral therapy can be competently done using one of the many commercially available encrypted video platforms. • Advanced techniques, such as virtual-reality procedures, can be effective in treating anxiety-related disorders during quarantine. The general population has experienced a significant elevation in fear and anxiety during COVID-19 both as a direct result of the virus but also due to measures taken to prevent it spreading, such as the need to stay inside and increase hand-washing. Lockdown has been used in many/most countries to prevent widespread infection. The advice and imposed actions are necessary to prevent the virus from spreading, but they might exacerbate the problems experienced by people with a preexisting anxiety-related disorder. The treatment of anxiety-related disorders can be provided while in quarantine. Staying at home in self-isolation does not preclude obtaining psychological treatment for anxiety-related disorders. Dealing with cognitive biases, over-estimations of threat, intolerance of uncertainty, inflated responsibility and excessive safety behavior, are useful clinical directions. [ABSTRACT FROM AUTHOR]

As part of a programme examining the effectiveness of land tenure forms − tenure administration system combinations, the paper examines how and why beneficiaries use land registration in Project 2, a state subsidised housing project in Mbekweni, South Africa. The ownership − registration combination can have a number of negative consequences when policy makers focus on economics alone, rather than the complete set of factors that are critical for registered ownership to work in poor urban communities. South Africa’s state-subsidised housing programme has created up to 3.7 million housing opportunities. Off-register secondary transactions constitute a major problem when the properties registered in ownership. Building on a series of related case studies, an evaluative schema classifying an ownership − registration combination as strong, semi-strong, semi-weak or weak is presented, along with the main findings from three previously reported case studies where the combination was classified as semi-strong. The beneficiaries in Project 2 were predominantly in-migrants from a rural customary area, and based on earlier work and the literature, a weak or semi-weak classification was expected. However, the case was classified as semi-strong. Ongoing, visible administration is possibly a major factor underlying this classification. [ABSTRACT FROM AUTHOR]

Abstract: This study tested Rachman''s cognitive behavioral method for treating obsessions not accompanied by prominent overt compulsions. The cognitive behavioral treatment was compared to waitlist control and an active and credible comparison of stress management training (SMT). Of the 73 adults who were randomized, 67 completed treatment, and 58 were available for one-year follow-up. The active treatments, compared to waitlist, resulted in substantially lower YBOCS scores, OCD-related cognitions and depression as well as improved social functioning. Overall, CBT and SMT showed large and similar reductions in symptoms. Pre–post effect sizes on YBOCS Obsessions for CBT and SMT completers was d = 2.34 and 1.90, respectively. Although CBT showed small advantages over SMT on some symptom measures immediately after treatment, these differences were no longer apparent in the follow-up period. CBT resulted in larger changes on most OCD-related cognitions compared to SMT. The cognitive changes were stable at 12 months follow-up, but the differences in the cognitive measures faded. The robust and enduring effects of both treatments contradict the long-standing belief that obsessions are resistant to treatment. [Copyright &y& Elsevier]

Abstract: Cognitive-behavioral therapies for anxiety disorders are highly efficacious (e.g., Butler, Chapman, Forman, & Beck, 2006; Deacon & Abramowitz, 2004). These treatments nevertheless remain underutilized and difficult to access for many of the patients who suffer from these conditions (e.g., Norton & Hope, 2005). We identify various barriers to the wide-scale dissemination of these treatments, including those that are applicable to empirically supported treatments more generally (e.g., lack of training opportunities, failure to address practitioner concerns) as well as those that may be relatively specific to CBT for anxiety disorders (e.g., practitioner concerns around using exposure interventions). We offer suggestions for overcoming these barriers, including specific guidance about continued accumulation of a supportive research base, making the appeals that are necessary to obtain required funding and organizational support, and the training of practitioners to deliver these treatments. Advocates of CBT for anxiety disorders will need to demonstrate that these treatments are cost effective, if wide-scale dissemination is to occur. In the United States, advocacy with third party payers will also be necessary. Although providing such steps may prove to be a difficult endeavour, the patients who stand to benefit from this work deserve nothing less. [Copyright &y& Elsevier]

Abstract: Contemporary cognitive treatment of obsessive-compulsive disorder (OCD) dates back to 1985, and rests on the premise that infrequent unwanted intrusions are essentially universal. As such, it is not the intrusion that is the focus of treatment but rather the interpretation or appraisal placed upon the intrusion. A number of cognitive domains are thought to be key areas, including inflated responsibility, overimportance of thoughts, need to control thoughts and overestimation of threat. Cognitive treatment of OCD is idiographic owing in part to the heterogeneous nature of the disorder as well as the cognitive domains implicated in the etiology and maintenance of the disorder. Cognitive treatment of an individual with ego-dystonic sexual and aggressive intrusions is illustrated in the current manuscript. Video clips are embedded at key moments during the treatment to ideally facilitate the illustration of this complex treatment. It is hoped that the addition of video clips will aid in the dissemination of this treatment that was recently tested and found to produce large effect sizes in a recent randomized controlled trial. [Copyright &y& Elsevier]

Naphthenic acids, with the general formula C n H2n + Z O2, are found in conventional petroleums and oil sands ores. These acids are toxic to aquatic life, so their discharge from petroleum processing into receiving waters must be avoided. In a previous study, naphthenic acids were putatively identified in groundwaters from two domestic wells that were distant from petroleum sources. However, coal deposits were near these wells. In this study, waters from the two wells were extracted and analyzed by electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry to unequivocally confirm the presence of naphthenic acids and other organic acids. In addition, distilled water was percolated through three crushed coal samples and the leachates were shown to contain a variety of organic acids, including naphthenic acids. These results clearly demonstrate that coal is a source of naphthenic acids and that the naphthenic acids can leach into groundwaters. Thus, the presence of naphthenic acids in waters cannot be solely attributed to petroleum or petroleum industry activities. [Copyright &y& Elsevier]

Abstract: This article presents the case of a 51-year old woman with obsessive-compulsive disorder. “Caroline” reported obsessions of harming people secondary to spreading her “bad energy,” which is experienced as dust on her hands and in her mouth. To prevent harm coming to others she mentally “vacuums” the dust, creates mental protective barriers around nearby people and avoids touching others for fear of transmitting her bad energy. Although she reported a childhood onset of obsessions and compulsions, it exacerbated in the context of multiple stressors 4 years prior to seeking treatment. Her Yale-Brown Obsessive Compulsive Scale total score was in the moderate range at intake. Predisposing, precipitating, and maintaining factors are described in addition to the specific symptom presentation. [Copyright &y& Elsevier]

The steroidogenic acute regulatory protein (StAR) is required for adrenal and gonadal steroidogenesis and for male sexual differentiation. StAR acts on the outer mitochondrial membrane (OMM) to facilitate movement of cholesterol from the OMM to the inner mitochondrial membrane to be converted to pregnenolone, the precursor of all steroid hormones. The mechanisms of the action of StAR remain unclear; the peripheral benzodiazepine receptor, an OMM protein, appears to be involved, but the identity of OMM proteins that interact with StAR remain unknown. Here we demonstrate that phosphorylated StAR interacts with voltage-dependent anion channel 1 (VDAC1) on the OMM, which then facilitates processing of the 37-kDa phospho-StAR to the 32-kDa intermediate. In the absence of VDAC1, phospho-StAR is degraded by cysteine proteases prior to mitochondrial import. Phosphorylation of StAR by protein kinase A requires phosphate carrier protein on the OMM, which appears to interact with StAR before it interacts with VDAC1. VDAC1 and phosphate carrier protein are the first OMM proteins shown to contact StAR. [ABSTRACT FROM AUTHOR]

Abstract: The efficacy of contemporary cognitive therapy for obsessive–compulsive disorder (OCD) has only recently been investigated. The current study compares exposure and response prevention (ERP) and cognitive behavior therapy (CBT) delivered in an individual format. Participants were randomly assigned to the 12 consecutive-week CBT or ERP treatment. Based on 59 treatment completers, there was no significant difference in YBOCS scores between CBT and ERP at post-treatment or at 3-month follow-up. A higher percentage of CBT participants obtained recovered status at post-treatment (67%) and at follow-up (76%), compared to ERP participants (59% and 58%, respectively), but the difference was not significant. Effect sizes (ESs) were used to compare the results of the current study with a previous study conducted at our center that utilized group CBT and ERP treatments, as well as other controlled trials that have compared CBT and ERP. The significance of these results is discussed and a comparison is made with the existing literature. [Copyright &y& Elsevier]

Presents a study which aimed to determine if cognitive-behavior therapy is effective in assisting patients in their attempts to discontinue selective serotonin reuptake inhibitor (SSRI) treatment. Outcome of several outpatients with panic disorder and agoraphobia who discontinued their treatment with SSRI; Associated risks with the use of SSRI; Demographic and diagnostic information at intake on an individual subject basis.

Presents a meta-analysis of psychosocial and pharmacological treatments to bulimia nervosa. Binge and purge frequency; Self-reported and interviewer-rated depression; Self-reported eating attitudes; Medication and cognitive bahavioral therapy.

Investigates the relationship between panic expectancy, social demand and agoraphobic avoidance in a group of panic disorder subjects. Role of fear of social scrutiny in avoidance; Performance during behavioral avoidance task involving either low/high social challenge task; Prediction of panic made before entering the situation; Type of social challenge condition.

In rare diseases (RDs) studies, generic preference-based patient-reported outcome measures (PROMs) might not be collected. This study aims to systematically review published studies using a mapping approach to derive HSUVs from non-preference-based PROMs in RDs. [Extracted from the article]

There is increasing recognition that conventional appraisal approaches (e.g. cost/QALY) may be unsuitable for rare disease treatments. These special processes are characterised by: consideration of additional criteria, more leniency around quality of evidence, special reimbursement status, different requirements for economic models, greater disease-specific expert input, and/or special rare disease appraisal committees. [Extracted from the article]

Abstract: Given the high prevalence and enormous burden of mental disorders and the efficacy of CBT in reducing symptom severity of a number of acute disorders, it is reasonable to use these same CBT strategies at an earlier stage to prevent the full expression of emotional problems. In comparison to treatment outcome research, work in prevention of mental disorders is in its infancy. Ongoing and recent prevention trials for 4 Axis I problems are introduced and the challenges of doing this work will be addressed. The goal within each one of the articles is to provide concrete guidelines and examples of the clinical work done in each trial to facilitate therapeutic efforts for individual practitioners. [Copyright &y& Elsevier]

The inner mitochondrial membrane protein 3β-hydroxysteroid dehydrogenase 2 (3βHSD2) synthesizes progesterone and androstenedione through its dehydrogenase and isomerase activities. This bifunctionality requires 3βHSD2 to undergo a conformational change. Given its proximity to the proton pump, we hypothesized that pH influences 3βHSD2 conformation and thus activity. Circular dichroism (CD) showed that between pH 7.4 and 4.5, 3βHSD2 retained its primarily α-helical character with a decrease in α-helical content at lower pH values, whereas the β-sheet content remained unchanged throughout. Titrating the pH back to 7.4 restored the original conformation within 25 min. Metabolic conversion assays indicated peak 3βHSD2 activity at pH 4.5 with ∼2-fold more progesterone synthesized at pH 4.5 than at pH 3.5 and 7.4. Increasing the 3βHSD2 concentration from 1 to 40 μg resulted in a 7-fold increase in progesterone at pH 4.5, but no change at pH 7.4. Incubation with guanidinum hydrochloride (GdmHCl) showed a three-step cooperative unfolding of 3βHSD2 from pH 7.4 to 4.5, possibly due to the native state unfolding to the intermediate ion core state. With further decreases in pH, increasing concentrations of GdmHCl led to rapid two-step unfolding that may represent complete loss of structure. Between pH 4 and 5, the two intermediate states appeared stable. Stopped-flow kinetics showed slower unfolding at around pH 4, where the protein is in a pseudostable state. Based on our data, we conclude that at pH 4-5, 3βHSD2 takes on a molten globule conformation that promotes the dual functionality of the enzyme. [ABSTRACT FROM AUTHOR]

Abstract: There is increasing international interest in naphthenic acids (NAs, classical formula CnH2n+ZO2) found in the oil sands from Alberta, Canada and in petroleum from around the world. The complexity of NAs poses major analytical challenges for their quantification and characterization. We used ultrahigh resolution electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (ESI-FT-ICR MS) to probe the make up of NAs from various sources by searching for peaks corresponding to the formula CnH2n+ZOx, for combinations of n=8 to 30, Z=0 to −12, and x=2 to 5. The sources included three commercial NAs preparations, and the acid-extractable organics from eight oil sand process-affected waters (OSPW) and from six surface fresh waters. Extracts from OSPW contained between 1 and 7% sulfur. The mass spectra showed between 300 and 1880 peaks, with >99% of the peaks having m/z between 145 and 600. In most cases, <20% of the peaks were assigned as classical NAs (x=2) and oxy-NAs (x=3 to 5). The classical NAs from the OSPW were predominantly Z=−4 and −6, whereas those from the fresh waters were mainly Z=0, with palmitic and stearic acids being the major components in the fresh waters. Remarkably, when the peak abundances were considered, <50% of the total abundance could be assigned to the classical and oxy-NAs. Thus, >50% of the compounds in the extracts of OSPW were not “naphthenic acids”. Based on these findings, it appears that the term “naphthenic acids”, which has been used to describe the toxic extractable compounds in OSPW, should be replaced by a term such as "oil sands tailings water acid-extractable organics (OSTWAEO)". Classical and oxy-NAs are components of OSTWAEO, but this term would not be as misleading as “naphthenic acids”. [ABSTRACT FROM AUTHOR]

Edaravone is considered to be a potent antioxidant drug known to scavenge free radical species and prevent free radical-induced lipid peroxidation. In this study, we investigated the effect of edaravone on the myeloperoxidase (MPO) activity, an enzyme responsible for the production of an array of neutrophil-derived oxidants that can cause cellular damage. The addition of edaravone to the reaction of MPO and hydrogen peroxide (H 2 O 2) significantly enhanced the reduction of MPO Compound II back to native MPO. Interestingly, the MPO-mediated production of toxic hypochlorous acid exhibited a concentration-dependent biphasic effect, with the apparent optimal edaravone concentration at 10 μM. Oxidation of edaravone by MPO was examined by various analytical methods. An MPO-catalyzed product(s) of edaravone was identified at 350 nm by kinetic analysis of UV–Vis spectroscopy. Several MPO-catalyzed metabolites of edaravone were proposed from the LC-MS analyses, including oxidized dimers from edaravone radicals. Electron spin resonance (ESR) spin trapping detected a carbon-centred radical metabolite of edaravone. NMR studies revealed that there are two exchangeable hydrogens, one of which is on the α-carbon, justifying the carbon-centred edaravone radical produced from MPO. Despite the formation of an edaravone carbon-radical metabolite, it did not appear to effectively oxidize GSH (in comparison with phenoxyl radicals). Viability (ATP) and cytotoxicity (LDH release) assays showed a concentration-dependent effect of edaravone on HL-60 cells treated with either a bolus concentration of 30 μM H 2 O 2 or a flux of H 2 O 2 generated by 5 mM glucose and 10 mU/mL glucose oxidase. The H 2 O 2 -induced toxicity was ameliorated at high edaravone concentrations (100–200 μM). In contrast, low concentrations of edaravone (1–10 μM) exacerbated the H 2 O 2 -induced toxicity. However, the effect of edaravone at low concentration (0–10 μM) appeared more prominent with the LDH assay only. The cellular findings correlated with the biochemical studies with respect to hypochlorous acid formation. These findings provide interesting perspectives regarding the duality of edaravone as an antioxidant drug. Image 1 • Edaravone is a substrate for myeloperoxidase. • Edaravone (≤10 μM) induced HOCl formation but attenuated at ≥ 30 μM. • The edaravone free radical metabolite was characterized, and polymers were observed. • Edaravone increased or decreased H 2 O 2 cytotoxicity depending on its concentration. [ABSTRACT FROM AUTHOR]

The formation of isonicotinyl-nicotinamide adenine dinucleotide (INH-NAD+) by the mycobacterial catalase-peroxidase enzyme, KatG, was known to be the major component of the mode of action of isoniazid (INH), an anti-tuberculosis drug. However, there are other enzymes that may catalyze this reaction. We have previously reported that neutrophil myeloperoxidase (MPO) is capable of metabolizing INH through the formation of INH-NAD+ adduct, which could be attributed to being a possible mode of action of INH. However, eosinophilic infiltration of the lungs is more pronounced and characteristic of granulomas in Mycobacterium tuberculosis -infected patients. Thus, the aim of the present study is to investigate the role of eosinophil peroxidase (EPO), a key eosinophil enzyme, during INH metabolism and the formation of its active metabolite, INH-NAD+ using purified EPO and eosinophils isolated from asthmatic donors. UV–Vis spectroscopy revealed INH oxidation by EPO led to a new product (λ max = 326 nm) in the presence of NAD+. This adduct was confirmed to be INH-NAD+ using LC-MS analysis where the intact adduct was detected (m / z = 769). Furthermore, EPO catalyzed the oxidation of INH and formed several free radical intermediates as assessed by electron paramagnetic resonance (EPR) spin-trapping; a carbon-centred radical, which is considered to be the reactive metabolite that binds with NAD+, was found when superoxide dismutase was included in the reaction. Our findings suggest that eosinophilic EPO may also play a role in the pharmacological activity of INH through the formation of INH-NAD+ adduct, and supports further evidence that human cells and enzymes are capable of producing the active metabolite involved in tuberculosis treatment. Image 1 • Eosinophil peroxidase (EPO) oxidizes isoniazid (INH). • Purified EPO formed a new product between INH and NAD+. • A carbon radical, formed by catalytic oxidation of INH by EPO, binds with NAD+. • EPO may contribute to the activity of INH by forming INH-NAD+ adduct. [ABSTRACT FROM AUTHOR]

Over the past 25 years Mental Contamination (MC) has become recognised as a distinct construct, particularly in relation to Obsessive Compulsive Disorder (OCD). MC is defined as feelings of contamination, often located internally, that arise in the absence of contact with a contaminant, with the source proposed to be human. Despite considerable interest from researchers and clinicians, there has not been a systematic review on the relationship between MC and OCD. Therefore, a systematic review was conducted to summarise and synthesise the current status of phenomenological and experimental evidence, mechanisms, assessment, measurement, and treatment of MC in OCD (PROSPERO: CRD42021223119). All study designs were eligible provided the focus of the study was on MC and the implications of the study were linked to OCD. We searched PsychINFO, Embase, Medline, Ethos, ProQuest, conference abstracts and trial registries between 1990 and 2021. The Mixed Methods Appraisal tool was used to assess methodological quality of included studies. We found 58 reports with a total of 67 studies that met criteria for inclusion in the review. Twenty-three of these studies used clinical samples, 28 were experimental, 12 focused on phenomenology and 8 addressed treatment. The quality of the studies was variable. Grey literature was not included, thus there may be further unpublished MC studies that have not been included in the review. Conclusions: Based on the findings, mental contamination is a robust clinical construct within OCD that has important implications for understanding and treating the disorder. • A systematic review examined the role MC in OCD. • We reviewed how MC in OCD is currently assessed, measured and treated. • A total of 58 reports which comprised 67 studies were included in the review. • The review findings indicate that MC is a distinct construct. • The relationship between MC and OCD has implications for assessment and treatment. [ABSTRACT FROM AUTHOR]

The formation of isonicotinyl-nicotinamide adenine dinucleotide (INH-NAD + ) via the mycobacterial catalase-peroxidase enzyme, KatG, has been described as the major component of the mode of action of isoniazid (INH). However, there are numerous human peroxidases that may catalyze this reaction. The role of neutrophil myeloperoxidase (MPO) in INH-NAD + adduct formation has never been explored; this is important, as neutrophils are recruited at the site of tuberculosis infection (granuloma) through infected macrophages’ cell death signals. In our studies, we showed that neutrophil MPO is capable of INH metabolism using electron paramagnetic resonance (EPR) spin-trapping and UV–Vis spectroscopy. MPO or activated human neutrophils (by phorbol myristate acetate) catalyzed the oxidation of INH and formed several free radical intermediates; the inclusion of superoxide dismutase revealed a carbon-centered radical which is considered to be the reactive metabolite that binds with NAD + . Other human metabolites, including N-acetyl-INH, N-acetylhydrazine, and hydrazine did not show formation of carbon-centered radicals, and either produced no detectable free radicals, N-centered free radicals, or superoxide, respectively. A comparison of these free radical products indicated that only the carbon-centered radical from INH is reducing in nature, based on UV–Vis measurement of nitroblue tetrazolium reduction. Furthermore, only INH oxidation by MPO led to a new product ( λ max = 326 nm) in the presence of NAD + . This adduct was confirmed to be isonicotinyl-NAD + using LC–MS analysis where the intact adduct was detected ( m / z = 769). The findings of this study suggest that neutrophil MPO may also play a role in INH pharmacological activity. [ABSTRACT FROM AUTHOR]

Steroid hormones are essential for carbohydrate metabolism, stress management, and reproduction and are synthesized from cholesterol in mitochondria of adrenal glands and gonads/ovaries. In acute stress or hormonal stimulation, steroidogenic acute regulatory protein (StAR) transports substrate cholesterol into the mitochondria for steroidogenesis by an unknown mechanism. Here, we report for the first time that StAR interacts with voltage-dependent anion channel 2 (VDAC2) at the mitochondria- associated endoplasmic reticulum membrane (MAM) prior to its translocation to the mitochondrial matrix. In the MAM, StAR interacts with mitochondrial proteins Tom22 and VDAC2. However, Tom22 knockdown by siRNA had no effect on pregnenolone synthesis. In the absence of VDAC2, StAR was expressed but not processed into the mitochondria as a mature 30-kDa protein. VDAC2 interacted with StAR via its C-terminal 20 amino acids and N-terminal amino acids 221-229, regulating the mitochondrial processing of StAR into the mature protein. In the absence of VDAC2, StAR could not enter the mitochondria or interact with MAM-associated proteins, and therefore steroidogenesis was inhibited. Furthermore, the N terminus was not essential for StAR activity, and the N-terminal deletion mutant continued to interact with VDAC2. The endoplasmic reticulum-targeting prolactin signal sequence did not affect StAR association with the MAM and thus its mitochondrial targeting. Therefore, VDAC2 controls StAR processing and activity, and MAM is thus a central location for initiating mitochondrial steroidogenesis. [ABSTRACT FROM AUTHOR]

In the adrenals, testes, and ovaries, 3β-hydroxysteroid dehydrogenase type 2 (3βHSD2) catalyzes the conversion of pregnenolone to progesterone and dehydroepiandrostenedione to androstenedione. Alterations in this pathway can have deleterious effects, including sexual development impairment, spontaneous abortion, and breast cancer. 3βHSD2, synthesized in the cytosol, is imported into the inner mitochondrial membrane (IMM) by translocases. Steroidogenesis requires that 3βHSD2 acts as both a dehydrogenase and isomerase. To achieve this dual functionality, 3βHSD2 must undergo a conformational change; however, what triggers that change remains unknown. We propose that 3βHSD2 associates with IMM or outer mitochondrial membrane translocases facing the intermembrane space (IMS) and that this interaction promotes the conformational change needed for full activity. Fractionation assays demonstrate that 3βHSD2 associated with the IMM but did not integrate into the membrane. Through mass spectrometry and Western blotting of mitochondrial complexes and density gradient ultracentrifugation, we show that that 3βHSD2 formed a transient association with the translocases Tim50 and Tom22 and with Tim23. This association occurred primarily through the interaction of Tim50 with the N terminus of 3βHSD2 and contributed to enzymatic activity. Tim50 knockdown inhibited catalysis of dehydroepiandrostenedione to androstenedione and pregnenolone to progesterone. Although Tim50 knockdown decreased 3βHSD2 expression, restoration of expression via proteasome and protease inhibition did not rescue activity. In addition, protein fingerprinting and CD spectroscopy reveal the flexibility of 3βHSD2, a necessary characteristic for forming multiple associations. In summary, Tim50 regulates 3βHSD2 expression and activity, representing a new role for translocases in steroidogenesis. [ABSTRACT FROM AUTHOR]

Glycoproteins constitute a class of compounds of increasing importance for pharmaceutical applications. The manipulation of bacterial protein glycosylation systems from Gram negative bacteria for the synthesis of recombinant glycoproteins is a promising alternative to the current production methods. Proteins carrying Lewis antigens have been shown to have potential applications for the treatment of diverse autoimmune diseases. In this work, we developed a mixed approach consisting of in vivo and in vitro steps for the synthesis of glycoproteins containing the Lewis X antigen. Using glycosyltransferases from Haemophilus influenzae, we engineered Escherichia coli to assemble a tetrasaccharide on the lipid carrier undecaprenylphosphate. This glycan was transferred in vivo from the lipid to a carrier protein by the Campylobacter jejuni oligosaccharyltransferase PglB. The glycoprotein was then fucosylated in vitro by a truncated fucosyltransferase from Helicobacter pylori. Diverse mass spectrometry techniques were used to confirm the structure of the glycan. The strategy presented here could be adapted in the future for the synthesis of diverse glycoproteins. Our experiments demonstrate that bacterial enzymes can be exploited for the production of glycoproteins carrying glycans present in human cells for potential therapeutic applications. [ABSTRACT FROM AUTHOR]