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Improved 2α-Hydroxylation Efficiency of Steroids by CYP154C2 Using Structure-Guided Rational Design.Applied and Environmental Microbiology Mar 2023Cytochrome P450 enzymes are promising biocatalysts for industrial use because they catalyze site-selective C-H oxidation and have diverse catalytic reactions and a broad...
Cytochrome P450 enzymes are promising biocatalysts for industrial use because they catalyze site-selective C-H oxidation and have diverse catalytic reactions and a broad substrate range. In this study, the 2α-hydroxylation activity of CYP154C2 from Streptomyces avermitilis MA-4680T toward androstenedione (ASD) was identified by an conversion assay. The testosterone (TES)-bound structure of CYP154C2 was solved at 1.42 Å, and this structure was used to design eight mutants, including single, double, and triple mutants, to improve the conversion efficiency. Mutants L88F/M191F and M191F/V285L were found to enhance the conversion rates significantly (i.e., 8.9-fold and 7.4-fold for TES, 46.5-fold and 19.5-fold for ASD, respectively) compared with the wild-type (WT) enzyme while retaining high 2α-position selectivity. The substrate binding affinity of the L88F/M191F mutant toward TES and ASD was enhanced compared with that of WT CYP154C2, supporting the measured increase in the conversion efficiencies. Moreover, the total turnover number and / of the L88F/M191F and M191F/V285L mutants increased significantly. Interestingly, all mutants containing L88F generated 16α-hydroxylation products, suggesting that L88 in CYP154C2 plays a vital role in substrate selectivity and that the amino acid corresponding to L88 in the 154C subfamily affects the orientation of steroid binding and substrate selectivity. Hydroxylated derivatives of steroids play essential roles in medicine. Cytochrome P450 enzymes selectively hydroxylate methyne groups on steroids, which can dramatically change their polarity, biological activity and toxicity. There is a paucity of reports on the 2α-hydroxylation of steroids, and documented 2α-hydroxylate P450s show extremely low conversion efficiency and/or low regio- and stereoselectivity. This study conducted crystal structure analysis and structure-guided rational engineering of CYP154C2 and efficiently enhanced the conversion efficiency of TES and ASD with high regio- and stereoselectivity. Our results provide an effective strategy and theoretical basis for the 2α-hydroxylation of steroids, and the structure-guided rational design of P450s should facilitate P450 applications in the biosynthesis of steroid drugs.
Topics: Hydroxylation; Steroids; Cytochrome P-450 Enzyme System; Oxidation-Reduction; Testosterone; Substrate Specificity
PubMed: 36847541
DOI: 10.1128/aem.02186-22 -
Harm Reduction Journal Dec 2019A growing body of evidence suggests that anabolic androgenic steroids (AAS) are used globally by a diverse population with varying motivations. Evidence has increased... (Review)
Review
BACKGROUND
A growing body of evidence suggests that anabolic androgenic steroids (AAS) are used globally by a diverse population with varying motivations. Evidence has increased greatly in recent years to support understanding of this form of substance use and the associated health harms, but there remains little evidence regarding interventions to support cessation and treat the consequences of use. In this scoping review, we identify and describe what is known about interventions that aim to support and achieve cessation of AAS, and treat and prevent associated health problems.
METHODS
A comprehensive search strategy was developed in four bibliographic databases, supported by an iterative citation searching process to identify eligible studies. Studies of any psychological or medical treatment interventions delivered in response to non-prescribed use of AAS or an associated harm in any setting were eligible.
RESULTS
In total, 109 eligible studies were identified, which included case reports representing a diverse range of disciplines and sources. Studies predominantly focussed on treatments for harms associated with AAS use, with scant evidence on interventions to support cessation of AAS use or responding to dependence. The types of conditions requiring treatment included psychiatric, neuroendocrine, hepatic, kidney, cardiovascular, musculoskeletal and infectious. There was limited evidence of engagement with users or delivery of psychosocial interventions as part of treatment for any condition, and of harm reduction interventions initiated alongside, or following, treatment. Findings were limited throughout by the case report study designs and limited information was provided.
CONCLUSION
This scoping review indicates that while a range of case reports describe treatments provided to AAS users, there is scarce evidence on treating dependence, managing withdrawal, or initiating behaviour change in users in any settings. Evidence is urgently required to support the development of effective services for users and of evidence-based guidance and interventions to respond to users in a range of healthcare settings. More consistent reporting in articles of whether engagement or assessment relating to AAS was initiated, and publication within broader health- or drug-related journals, will support development of the evidence base.
Topics: Delivery of Health Care; Harm Reduction; Humans; Psychotherapy; Social Support; Substance-Related Disorders; Testosterone Congeners
PubMed: 31888665
DOI: 10.1186/s12954-019-0343-1 -
Journal of the American Academy of... Jan 2022Despite the well-documented effects of testosterone and its synthetic derivatives-collectively termed anabolic androgenic steroids (AASs)-on the musculoskeletal system,... (Review)
Review
Despite the well-documented effects of testosterone and its synthetic derivatives-collectively termed anabolic androgenic steroids (AASs)-on the musculoskeletal system, the therapeutic use of these agents has received limited investigation within the field of orthopaedic surgery. In the last 2 decades, preclinical and clinical research has started to identify promising applications of the short-term use of AASs in the perioperative period. There is evidence to suggest that AASs may improve postoperative recovery after anterior cruciate ligament reconstruction and total joint arthroplasty. In addition, AASs may augment the biological healing environment in specific clinical scenarios including muscle injury, fracture repair, and rotator cuff repair. Current literature fails to present strong evidence for or against the use of AASs in orthopaedics, but there is continuous research on this topic. The purpose of this study was to provide a comprehensive overview of the current status of AAS applications in orthopaedic surgery, with an emphasis on preclinical data, clinical studies, and future directions.
Topics: Anabolic Agents; Humans; Orthopedic Procedures; Orthopedics; Steroids; Testosterone Congeners
PubMed: 34982051
DOI: 10.5435/JAAOSGlobal-D-21-00156 -
International Journal of Molecular... Apr 2016Dietary supplements (DS) are extensively consumed worldwide despite unproven efficacy. The true incidence of DS-induced liver injury (DSILI) is unknown but is probably... (Review)
Review
Dietary supplements (DS) are extensively consumed worldwide despite unproven efficacy. The true incidence of DS-induced liver injury (DSILI) is unknown but is probably under-diagnosed due to the general belief of safety of these products. Reported cases of herbals and DS-induced liver injury are increasing worldwide. The aim of this manuscript is to report a tabular listing with a description of DS associated with hepatotoxicity as well as review the phenotype and severity of DSILI. Natural remedies related to hepatotoxicity can be divided into herbal product-induced liver injury and DS-induced liver injury. In this article, we describe different DS associated with liver injury, some of them manufactured DS containing several ingredients (Herbalife™ products, Hydroxycut™, LipoKinetix™, UCP-1 and OxyELITE™) while others have a single ingredient (green tea extract, linoleic acid, usnic acid, 1,3-Dimethylamylamine, vitamin A, Garcinia cambogia and ma huang). Additional DS containing some of the aforementioned ingredients implicated in liver injury are also covered. We have also included illicit androgenic anabolic steroids for bodybuilding in this work, as they are frequently sold under the denomination of DS despite being conventional drugs.
Topics: Chemical and Drug Induced Liver Injury; Clinical Trials as Topic; Dietary Supplements; Female; Humans; Male; Plant Preparations; Plants, Medicinal; Testosterone Congeners
PubMed: 27070596
DOI: 10.3390/ijms17040537 -
Neurochemistry International 2008Dehydroepiandrosterone (DHEA) is an abundant circulating prohormone in humans, with a variety of reported actions on central and peripheral tissues. Despite its... (Review)
Review
Dehydroepiandrosterone (DHEA) is an abundant circulating prohormone in humans, with a variety of reported actions on central and peripheral tissues. Despite its abundance, the functions of DHEA are relatively unknown because common animal models (laboratory rats and mice) have very low DHEA levels in the blood. Over the past decade, we have obtained considerable evidence from avian studies demonstrating that (1) DHEA is an important circulating prohormone in songbirds and (2) the enzyme 3beta-hydroxysteroid dehydrogenase/isomerase (3beta-HSD), responsible for converting DHEA into a more active androgen, is expressed at high levels in the songbird brain. Here, we first review biochemical and molecular studies demonstrating the widespread activity and expression of 3beta-HSD in the adult and developing songbird brain. Studies examining neural 3beta-HSD activity show effects of sex, stress, and season that are region-specific. Second, we review studies showing seasonal and stress-related changes in circulating DHEA in captive and wild songbird species. Third, we describe evidence that DHEA treatment can stimulate song behavior and the growth of neural circuits controlling song behavior. Importantly, brain 3beta-HSD and aromatase can work in concert to locally metabolize DHEA into active androgens and estrogens, which are critical for controlling behavior and robust adult neuroplasticity in songbirds. DHEA is likely secreted by the avian gonads and/or adrenals, as is the case in humans, but DHEA may also be synthesized de novo in the songbird brain from cholesterol or other precursors. Irrespective of its source, DHEA seems to be an important prohormone in songbirds, and 3beta-HSD is a key enzyme in the songbird brain.
Topics: 3-Hydroxysteroid Dehydrogenases; Animals; Behavior, Animal; Biotransformation; Brain; Brain Chemistry; Cells, Cultured; Dehydroepiandrosterone; Gonadal Steroid Hormones; Songbirds
PubMed: 17643555
DOI: 10.1016/j.neuint.2007.05.003 -
Sensors (Basel, Switzerland) Dec 2021Anabolic-androgenic steroids (AASs), a group of compounds frequently misused by athletes and, unfortunately, also by the general population, have lately attracted global... (Review)
Review
Anabolic-androgenic steroids (AASs), a group of compounds frequently misused by athletes and, unfortunately, also by the general population, have lately attracted global attention; thus, significant demands for more precise, facile, and rapid AAS detection have arisen. The standard methods ordinarily used for AAS determination include liquid and gas chromatography coupled with mass spectrometry. However, good knowledge of steroid metabolism, pretreatment of samples (such as derivatization), and well-trained operators of the instruments are required, making this procedure expensive, complicated, and not routinely applicable. In the drive to meet current AAS detection demands, the scientific focus has shifted to developing novel, tailor-made approaches leading to time- and cost-effective, routine, and field-portable methods for AAS determination in various matrices, such as biological fluids, food supplements, meat, water, or other environmental components. Therefore, herein, we present a comprehensive review article covering recent advances in AAS determination, with a strong emphasis on the increasingly important role of chemically designed artificial sensors, biosensors, and antibody- and fluorescence-based methods.
Topics: Anabolic Agents; Androgens; Athletes; Doping in Sports; Humans; Steroids; Testosterone Congeners
PubMed: 35009549
DOI: 10.3390/s22010004 -
Frontiers in Endocrinology 2023Clinical studies have shown that low levels of endogenous testosterone are associated with cardiovascular diseases. Considering the intimate connection between oxidative...
INTRODUCTION
Clinical studies have shown that low levels of endogenous testosterone are associated with cardiovascular diseases. Considering the intimate connection between oxidative metabolism and myocardial contractility, we determined the effects of testosterone deficiency on the two spatially distinct subpopulations of cardiac mitochondria, subsarcolemmal (SSM) and interfibrillar (IFM).
METHODS
We assessed cardiac function and cardiac mitochondria structure of SSM and IFM after 12 weeks of testosterone deficiency in male Wistar rats.
RESULTS AND DISCUSSION
Results show that low testosterone reduced myocardial contractility. Orchidectomy increased total left ventricular mitochondrial protein in the SSM, but not in IFM. The membrane potential, size and internal complexity in the IFM after orchidectomy were higher compared to the SHAM group. However, the rate of oxidative phosphorylation with all substrates in the IFM after orchidectomy was lower compared to the SHAM group. Testosterone replacement restored these changes. In the testosterone-deficient SSM group, oxidative phosphorylation was decreased with palmitoyl-L-carnitine as substrate; however, the mitochondrial calcium retention capacity in IFM was increased. There was no difference in swelling of the mitochondria in either group. These changes in IFM were followed by a reduction in phosphorylated form of AMP-activated protein kinase (p-AMPK-α), peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) translocation to mitochondria and decreased mitochondrial transcription factor A (TFAM). Testosterone deficiency increased NADPH oxidase (NOX), angiotensin converting enzyme (ACE) protein expression and reduced mitochondrial antioxidant proteins such as manganese superoxide dismutase (Mn-SOD) and catalase in the IFM. Treatment with apocynin (1.5 mM in drinking water) normalized myocardial contractility and interfibrillar mitochondrial function in the testosterone depleted animals. In conclusion, our findings demonstrate that testosterone deficiency leads to reduced myocardial contractility and impaired cardiac interfibrillar mitochondrial function. Our data suggest the involvement of reactive oxygen species, with a possibility of NOX as an enzymatic source.
Topics: Rats; Animals; Male; Rats, Wistar; Myocardium; Mitochondria, Heart; Oxidative Stress; Testosterone
PubMed: 37780627
DOI: 10.3389/fendo.2023.1206387 -
Journal of Andrology 2009A considerable body of evidence exists suggesting that androgen deficiency contributes to the onset, progression, or both of cardiovascular disease (CVD). The aim of... (Review)
Review
A considerable body of evidence exists suggesting that androgen deficiency contributes to the onset, progression, or both of cardiovascular disease (CVD). The aim of this review is to evaluate the relationships between testosterone (T) deficiency and risk factors of CVD and to discuss the implications of androgen deficiency in men with cardiovascular risk factors. The relationship between androgen deficiency and endothelial function, lipid profiles, inflammatory responses, altered vascular smooth muscle reactivity, and hypertension are discussed with regard to CVD. A comprehensive literature search was carried out with the use of Pub Med from 1980 through 2009, and relevant articles pertinent to androgen deficiency and vascular disease were evaluated and discussed. Low T, whether attributed to hypogonadism or androgen deprivation therapy, in men with prostate carcinoma, produces adverse effects on cardiovascular health. Androgen deficiency is associated with increased levels of total cholesterol, low-density lipoprotein, increased production of proinflammatory factors, and increased thickness of the arterial wall and contributes to endothelial dysfunction. Testosterone supplementation restores arterial vasoreactivity; reduces proinflammatory cytokines, total cholesterol, and triglyceride levels; and improves endothelial function but also might reduce high-density lipoprotein levels. Testosterone is an anabolic hormone with a wide range of beneficial effects on men's health. The therapeutic role of T in men's health, however, remains a hotly debated issue for a number of reasons, including the purported risk of prostate cancer. In view of the emerging evidence suggesting that androgen deficiency is a risk factor for CVD, androgen replacement therapy could potentially reduce CVD risk in hypogonadal men. It should be emphasized, however, that androgen replacement therapy should be done with very thorough and careful monitoring for prostate diseases.
Topics: Animals; Cardiovascular Diseases; Humans; Hypertension; Hypogonadism; Lipid Metabolism; Male; Metabolic Syndrome; Risk Factors; Testosterone; Vascular Diseases; Vasodilation
PubMed: 19342698
DOI: 10.2164/jandrol.108.007245 -
Journal of Investigative Medicine : the... Feb 2012The survival and progression of prostate cancer are generally dependent on expression of the androgen receptor (AR), as well as the availability of endogenous AR... (Review)
Review
The survival and progression of prostate cancer are generally dependent on expression of the androgen receptor (AR), as well as the availability of endogenous AR agonists. Originating from the gonads, testosterone is released into circulation and is converted by steroid-5α-reductase in prostate cancer to 5α-dihydrotestosterone (DHT), potently activating AR and driving tumor progression. Advanced prostate cancer is initially treated with gonadal testosterone depletion, which suppresses this cascade of events and typically leads to a treatment response. Eventually, resistance to testosterone deprivation occurs with "castration-resistant" prostate cancer (CRPC) and is driven by the intratumoral synthesis of DHT. The generation of DHT occurs in large part from adrenal 19-carbon precursor steroids, which are dependent on expression of CYP17A1. Although the path from adrenal precursor steroids to DHT was generally thought to require 5α-reduction of testosterone, recent data suggest that it instead involves conversion from Δ-androstenedione by steroid-5α-reductase isoenzyme-1 to 5α-androstanedione, followed by subsequent conversion to DHT. The 5α-androstanedione pathway to DHT therefore bypasses testosterone entirely. Abiraterone acetate effectively inhibits CYP17A1, blocks the synthesis of androgens, and extends the survival of men with CRPC. Further progress in the hormonal treatment of CRPC is dependent on an understanding of the mechanisms that underlie CRPC and resistance to abiraterone acetate.
Topics: Abiraterone Acetate; Androstadienes; Biosynthetic Pathways; Dihydrotestosterone; Etiocholanolone; Humans; Male; Orchiectomy; Prostatic Neoplasms
PubMed: 22064602
DOI: 10.2310/JIM.0b013e31823874a4 -
Neuroendocrinology 2012Anabolic androgenic steroids (AAS) comprise a large and growing class of synthetic androgens used clinically to promote tissue-building in individuals suffering from... (Review)
Review
Anabolic androgenic steroids (AAS) comprise a large and growing class of synthetic androgens used clinically to promote tissue-building in individuals suffering from genetic disorders, injuries, and diseases. Despite these beneficial therapeutic applications, the predominant use of AAS is illicit: these steroids are self-administered to promote athletic performance and body image. Hand in hand with the desired anabolic actions of the AAS are untoward effects on the brain and behavior. While the signaling routes by which the AAS impose both beneficial and harmful actions may be quite diverse, key endpoints are likely to include ligand-gated and voltage-dependent ion channels that govern the activity of electrically excitable tissues. Here, we review the known effects of AAS on molecular targets that play critical roles in controlling electrical activity, with a specific focus on the effects of AAS on neurotransmission mediated by GABA(A) receptors in the central nervous system.
Topics: Anabolic Agents; Animals; Brain; Humans; Ion Channels; Neurons; Receptors, GABA-A; Steroids; Synaptic Transmission; Testosterone Congeners
PubMed: 22576754
DOI: 10.1159/000339123