AANAT kinetics of CoASH-targeted electrophiles of tryptamine and related analogs.

Arylalkylamine N-acetyltransferase (AANAT) catalyzes the rate-limiting step in melatonin synthesis and is a potential target for disorders involving melatonin overproduction, such as seasonal affective disorder. Previously described AANAT inhibitor bromoacetyltryptamine (BAT) and benzothiophenes analogs were reported to react with CoASH to form potent bisubstrate inhibitors through AANAT's alkyltransferase function, which is secondary to its role as an acetyltransferase. We replaced the bromoacetyl group in BAT with various Michael acceptors to mitigate possible off-target activity of its bromoacetyl group.

Educational guidelines on sexual orientation, gender identity and expression, and sex characteristics biases in medical education.

A commitment to diversity, equity, inclusion, and belonging in medical education requires addressing both explicit and implicit biases based on sexual orientation, gender identity and expression, and sex characteristics and the intersectionality with other identities. Heterosexism and heteronormative attitudes contribute to health and healthcare disparities for lesbian, gay, bisexual, transgender and queer or questioning, intersex, asexual individuals. Student, trainee, and faculty competencies in medical education curricula regarding the care of lesbian, gay, bisexual, transgender and queer or questioning, intersex, asexual patients and those who are gender nonconforming or born with differences of sex development allow for better understanding and belonging within the clinical learning environment of lesbian, gay, bisexual, transgender and queer/questioning, intersex, asexual learners and educators.

Evaluation of Rhodanine Indolinones as AANAT Inhibitors.

Circadian rhythm (CR) dysregulation negatively impacts health and contributes to mental disorders. The role of melatonin, a hormone intricately linked to CR, is still a subject of active study. The enzyme arylalkylamine N-acetyltransferase (AANAT) is responsible for melatonin synthesis, and it is a potential target for disorders that involve abnormally high melatonin levels, such as seasonal affective disorder (SAD).

Educational guidelines for diversity and inclusion: addressing racism and eliminating biases in medical education.

Racism and bias contribute to healthcare disparities at a patient and population health level and also contribute to the stagnation or even regression of progress toward equitable representation in the workforce and in healthcare leadership. Medical education and healthcare systems have expended tremendous efforts over the past several years to address these inequities. However, systemic racism continues to impact health outcomes and the future physician workforce.

DJ-1 is not a deglycase and makes a modest contribution to cellular defense against methylglyoxal damage in neurons.

Human DJ-1 is a cytoprotective protein whose absence causes Parkinson's disease and is also associated with other diseases. DJ-1 has an established role as a redox-regulated protein that defends against oxidative stress and mitochondrial dysfunction. Multiple studies have suggested that DJ-1 is also a protein/nucleic acid deglycase that plays a key role in the repair of glycation damage caused by methylglyoxal (MG), a reactive α-keto aldehyde formed by central metabolism.

Factors Associated with Meeting Obstetric Care Consensus Guidelines for Nulliparous, Term, Singleton, Vertex Cesarean Births.

 This study aimed to identify factors associated with meeting the Obstetric Care Consensus (OCC) guidelines for nulliparous, term, singleton, and vertex (NTSV) cesarean births.  This was a retrospective case control study of women with NTSV cesarean births between January 2014 and December 2017 at single tertiary care center. Demographics and clinical characteristics were compared between women with NTSV cesarean births which did or did not meet OCC guidelines.

Detailed evaluation of pyruvate dehydrogenase complex inhibition in simulated exercise conditions.

The mammalian pyruvate dehydrogenase complex (PDC) is a mitochondrial multienzyme complex that connects glycolysis to the tricarboxylic acid cycle by catalyzing pyruvate oxidation to produce acetyl-CoA, NADH, and CO. This reaction is required to aerobically utilize glucose, a preferred metabolic fuel, and is composed of three core enzymes: pyruvate dehydrogenase (E1), dihydrolipoyl transacetylase (E2), and dihydrolipoyl dehydrogenase (E3). The pyruvate-dehydrogenase-specific kinase (PDK) and pyruvate-dehydrogenase-specific phosphatase (PDP) are considered the main control mechanism of mammalian PDC activity.

Computationally modeling mammalian succinate dehydrogenase kinetics identifies the origins and primary determinants of ROS production.

Succinate dehydrogenase (SDH) is an inner mitochondrial membrane protein complex that links the Krebs cycle to the electron transport system. It can produce significant amounts of superoxide ([Formula: see text]) and hydrogen peroxide (HO); however, the precise mechanisms are unknown. This fact hinders the development of next-generation antioxidant therapies targeting mitochondria.

Evidence of a preferred kinetic pathway in the carnitine acetyltransferase reaction.

Mammalian carnitine acetyltransferase (CrAT) is a mitochondrial enzyme that catalyzes the reversible transfer of an acetyl group from acetyl-CoA to carnitine. CrAT knockout studies have shown that this enzyme is critical to sustain metabolic flexibility, or the ability to switch between different fuel types, an underlying theme of the metabolic syndrome. These recent physiological findings imply that CrAT dysfunction, or its catalytic impairment, may lead to disease.

Systems-level computational modeling demonstrates fuel selection switching in high capacity running and low capacity running rats.

High capacity and low capacity running rats, HCR and LCR respectively, have been bred to represent two extremes of running endurance and have recently demonstrated disparities in fuel usage during transient aerobic exercise. HCR rats can maintain fatty acid (FA) utilization throughout the course of transient aerobic exercise whereas LCR rats rely predominantly on glucose utilization. We hypothesized that the difference between HCR and LCR fuel utilization could be explained by a difference in mitochondrial density.

Identification of a Conserved Histidine As Being Critical for the Catalytic Mechanism and Functional Switching of the Multifunctional Proline Utilization A Protein.

Proline utilization A from Escherichia coli (EcPutA) is a multifunctional flavoenzyme that oxidizes proline to glutamate through proline dehydrogenase (PRODH) and Δ-pyrroline-5-carboxylate dehydrogenase (P5CDH) activities, while also switching roles as a DNA-bound transcriptional repressor and a membrane-bound catabolic enzyme. This phenomenon, termed functional switching, occurs through a redox-mediated mechanism in which flavin reduction triggers a conformational change that increases EcPutA membrane binding affinity. Structural studies have shown that reduction of the FAD cofactor causes the ribityl moiety to undergo a crankshaft motion, indicating that the orientation of the ribityl chain is a key element of PutA functional switching.

Global Kinetic Analysis of Mammalian E3 Reveals pH-dependent NAD+/NADH Regulation, Physiological Kinetic Reversibility, and Catalytic Optimum.

Mammalian E3 is an essential mitochondrial enzyme responsible for catalyzing the terminal reaction in the oxidative catabolism of several metabolites. E3 is a key regulator of metabolic fuel selection as a component of the pyruvate dehydrogenase complex (PDHc). E3 regulates PDHc activity by altering the affinity of pyruvate dehydrogenase kinase, an inhibitor of the enzyme complex, through changes in reduction and acetylation state of lipoamide moieties set by the NAD(+)/NADH ratio.

A pH-dependent kinetic model of dihydrolipoamide dehydrogenase from multiple organisms.

Dihydrolipoamide dehydrogenase is a flavoenzyme that reversibly catalyzes the oxidation of reduced lipoyl substrates with the reduction of NAD(+) to NADH. In vivo, the dihydrolipoamide dehydrogenase component (E3) is associated with the pyruvate, α-ketoglutarate, and glycine dehydrogenase complexes. The pyruvate dehydrogenase (PDH) complex connects the glycolytic flux to the tricarboxylic acid cycle and is central to the regulation of primary metabolism.

Luminol-based chemiluminescent signals: clinical and non-clinical application and future uses.

Chemiluminescence (CL) is an important method for quantification and analysis of various macromolecules. A wide range of CL agents such as luminol, hydrogen peroxide, fluorescein, dioxetanes and derivatives of oxalate, and acridinium dyes are used according to their biological specificity and utility. This review describes the application of luminol chemiluminescence (LCL) in forensic, biomedical, and clinical sciences.

Evidence for hysteretic substrate channeling in the proline dehydrogenase and Δ1-pyrroline-5-carboxylate dehydrogenase coupled reaction of proline utilization A (PutA).

PutA (proline utilization A) is a large bifunctional flavoenzyme with proline dehydrogenase (PRODH) and Δ(1)-pyrroline-5-carboxylate dehydrogenase (P5CDH) domains that catalyze the oxidation of l-proline to l-glutamate in two successive reactions. In the PRODH active site, proline undergoes a two-electron oxidation to Δ(1)-pyrroline-5-carboxlylate, and the FAD cofactor is reduced. In the P5CDH active site, l-glutamate-γ-semialdehyde (the hydrolyzed form of Δ(1)-pyrroline-5-carboxylate) undergoes a two-electron oxidation in which a hydride is transferred to NAD(+)-producing NADH and glutamate.

The three-dimensional structural basis of type II hyperprolinemia.

Type II hyperprolinemia is an autosomal recessive disorder caused by a deficiency in Δ(1)-pyrroline-5-carboxylate dehydrogenase (P5CDH; also known as ALDH4A1), the aldehyde dehydrogenase that catalyzes the oxidation of glutamate semialdehyde to glutamate. Here, we report the first structure of human P5CDH (HsP5CDH) and investigate the impact of the hyperprolinemia-associated mutation of Ser352 to Leu on the structure and catalytic properties of the enzyme. The 2.

A role for aberrant protein palmitoylation in FFA-induced ER stress and β-cell death.

Exposure of insulin-producing cells to elevated levels of the free fatty acid (FFA) palmitate results in the loss of β-cell function and induction of apoptosis. The induction of endoplasmic reticulum (ER) stress is one mechanism proposed to be responsible for the loss of β-cell viability in response to palmitate treatment; however, the pathways responsible for the induction of ER stress by palmitate have yet to be determined. Protein palmitoylation is a major posttranslational modification that regulates protein localization, stability, and activity.

Rapid reaction kinetics of proline dehydrogenase in the multifunctional proline utilization A protein.

The multifunctional proline utilization A (PutA) flavoenzyme from Escherichia coli catalyzes the oxidation of proline to glutamate in two reaction steps using separate proline dehydrogenase (PRODH) and Δ(1)-pyrroline-5-carboxylate (P5C) dehydrogenase domains. Here, the kinetic mechanism of PRODH in PutA is studied by stopped-flow kinetics to determine microscopic rate constants for the proline:ubiquinone oxidoreductase mechanism. Stopped-flow data for proline reduction of the flavin cofactor (reductive half-reaction) and oxidation of reduced flavin by CoQ(1) (oxidative half-reaction) were best-fit by a double exponential from which maximum observable rate constants and apparent equilibrium dissociation constants were determined.

Steady-state kinetic mechanism of the proline:ubiquinone oxidoreductase activity of proline utilization A (PutA) from Escherichia coli.

The multifunctional proline utilization A (PutA) flavoenzyme from Escherichia coli performs the oxidation of proline to glutamate in two catalytic steps using separate proline dehydrogenase (PRODH) and Δ(1)-pyrroline-5-carboxylate (P5C) dehydrogenase domains. In the first reaction, the oxidation of proline is coupled to the reduction of ubiquinone (CoQ) by the PRODH domain, which has a β(8)α(8)-barrel structure that is conserved in bacterial and eukaryotic PRODH enzymes. The structural requirements of the benzoquinone moiety were examined by steady-state kinetics using CoQ analogs.

Flavin redox switching of protein functions.

Flavin cofactors impart remarkable catalytic diversity to enzymes, enabling them to participate in a broad array of biological processes. The properties of flavins also provide proteins with a versatile redox sensor that can be utilized for converting physiological signals such as cellular metabolism, light, and redox status into a unique functional output. The control of protein functions by the flavin redox state is important for transcriptional regulation, cell signaling pathways, and environmental adaptation.

Homogeneous insulin and C-Peptide sensors for rapid assessment of insulin and C-peptide secretion by the islets.

Objective: Glucose-stimulated islet insulin or C-peptide secretion experiments are a fundamental tool for studying and assessing islet function. The goal of this work was to develop Ab-based fluorescent homogenous sensors that would allow rapid, inexpensive, near-instantaneous determinations of insulin and C-peptide levels in biological samples.

Research Design And Methods: Our approach was to use molecular pincer design (Heyduk et al.

Marijuana use is not associated with cervical human papillomavirus natural history or cervical neoplasia in HIV-seropositive or HIV-seronegative women.

Marijuana use was recently reported to have a positive cross-sectional association with human papillomavirus (HPV)-related head and neck cancer. Laboratory data suggest that marijuana could have an immunomodulatory effect. Little is known, however, regarding the effects of marijuana use on cervical HPV or neoplasia.

Selective mtDNA mutation accumulation results in beta-cell apoptosis and diabetes development.

To test the hypothesis that somatic mitochondrial (mt)DNA mutation accumulation predisposes mice to beta-cell loss and diabetes development, transgenic mice expressing a proofreading-deficient mtDNA polymerase-gamma under the control of the rat insulin-1 promoter were generated. At 6 wk of age, mtDNA mutations reached 0.01% (1.

Insulin-like growth factor axis and oncogenic human papillomavirus natural history.

High serum levels of insulin-like growth factor-I (IGF-I) are reported to be a risk factor for several common cancers, and recent cross-sectional data suggest a possible additional association of IGF-I with cervical neoplasia. To prospectively assess whether circulating IGF-I levels influence the natural history of oncogenic human papillomavirus (HPV), the viral cause of cervical cancer, we conducted a pilot investigation of 137 women who underwent semiannual type-specific HPV DNA PCR testing and cervical cytology. Total IGF-I and IGF binding protein-3 (IGFBP-3), the most abundant IGFBP in circulation, were measured using baseline serum specimens.

Analysis of standard methods for diagnosing vaginitis: HIV infection does not complicate the diagnosis of vaginitis.

Objective: We sought to determine whether the standard diagnostic methods for vaginitis behave similarly among HIV-infected and at-risk seronegative women.

Materials And Methods: We performed pairwise comparisons over time (1994-2003) for the different diagnostic methods for bacterial vaginosis (BV) (Nugent score and Amsel criteria), vulvovaginal candidiasis (potassium hydroxide smear and Pap smear), and trichomoniasis (culture, wet mount, and Pap smear) among HIV-infected and at-risk HIV seronegative women in the Women's Interagency HIV Study cohort. We stratified subjects by HIV status and among the HIV-infected women by CD4+ cell count strata.