Cysteine hydropersulfide (CysSSH) occurs in abundant quantities in various organisms, yet little is known about its biosynthesis and physiological functions. Extensive persulfide formation is ...apparent in cysteine-containing proteins in Escherichia coli and mammalian cells and is believed to result from post-translational processes involving hydrogen sulfide-related chemistry. Here we demonstrate effective CysSSH synthesis from the substrate L-cysteine, a reaction catalyzed by prokaryotic and mammalian cysteinyl-tRNA synthetases (CARSs). Targeted disruption of the genes encoding mitochondrial CARSs in mice and human cells shows that CARSs have a crucial role in endogenous CysSSH production and suggests that these enzymes serve as the principal cysteine persulfide synthases in vivo. CARSs also catalyze co-translational cysteine polysulfidation and are involved in the regulation of mitochondrial biogenesis and bioenergetics. Investigating CARS-dependent persulfide production may thus clarify aberrant redox signaling in physiological and pathophysiological conditions, and suggest therapeutic targets based on oxidative stress and mitochondrial dysfunction.
Abstract Background Postoperative adhesions are a significant health problem with major implications on quality of life and health care expenses. The purpose of this review was to investigate the ...efficacy of preventative techniques and adhesion barriers and identify those patients who are most likely to benefit from these strategies. Methods The National Library of Medicine, Medline, Embase, and Cochrane databases were used to identify articles related to postoperative adhesions. Results Ileal pouch–anal anastomosis, open colectomy, and open gynecologic procedures are associated with the highest risk of adhesive small-bowel obstruction (class I evidence). Based on expert opinion (class III evidence) intraoperative preventative principles, such as meticulous hemostasis, avoiding excessive tissue dissection and ischemia, and reducing remaining surgical material have been published. Laparoscopic techniques, with the exception of appendicitis, result in fewer adhesions than open techniques (class I evidence). Available bioabsorbable barriers, such as hyaluronic acid/carboxymethylcellulose and icodextrin 4% solution, have been shown to reduce adhesions (class I evidence). Conclusions Postoperative adhesions are a significant health problem with major implications on quality of life and health care. General intraoperative preventative techniques, laparoscopic techniques, and the use of bioabsorbable mechanical barriers in the appropriate cases reduce the incidence and severity of peritoneal adhesions.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
ERGIC-53 transports certain subsets of newly synthesized secretory proteins and membrane proteins from the endoplasmic reticulum to the Golgi apparatus. Despite numerous structural and functional ...studies since its identification, the overall architecture and mechanism of action of ERGIC-53 remain unclear. Here we present cryo-EM structures of full-length ERGIC-53 in complex with its functional partner MCFD2. These structures reveal that ERGIC-53 exists as a homotetramer, not a homohexamer as previously suggested, and comprises a four-leaf clover-like head and a long stalk composed of three sets of four-helix coiled-coil followed by a transmembrane domain. 3D variability analysis visualizes the flexible motion of the long stalk and local plasticity of the head region. Notably, MCFD2 is shown to possess a Zn
-binding site in its N-terminal lid, which appears to modulate cargo binding. Altogether, distinct mechanisms of cargo capture and release by ERGIC- 53 via the stalk bending and metal binding are proposed.
Background
Since 2007, clinical practice guidelines by the Society of American Gastrointestinal and Endoscopic Surgeons (SAGES) recommend early surgical management with laparoscopic cholecystectomy ...for pregnant women with symptomatic gallbladder disease regardless of trimester. However, little is known about practice patterns in the management of pregnant patients with acute cholecystitis. This study aims to examine nationwide trends in the surgical management of acute cholecystitis, as well as their impact on clinical outcomes during pregnancy.
Methods
The National Inpatient Sample was queried for all pregnant women diagnosed with acute cholecystitis between January 2003 and September 2015. After applying appropriate weights, multivariate regression analysis adjusted for patient- and hospital-level characteristics and quantified the impact of discharge year (2003–2007 versus 2008–2015) on cholecystectomy rates and timing of surgery. Multivariate regression analysis was also used to examine the impact of same admission cholecystectomy and its timing on maternal and fetal outcomes.
Results
A total of 23,939 pregnant women with acute cholecystitis satisfied our inclusion criteria. The median age was 26 years (interquartile range: 22–30). During the study period, 36.3% were managed non-operatively while 59.6% and 4.1% underwent laparoscopic and open cholecystectomy, respectively. After adjusting for covariates, laparoscopic cholecystectomy was more commonly performed after 2007 (odds ratio OR 1.333,
p
< 0.001). Furthermore, time from admission to surgery was significantly shorter in the latter study period (regression coefficient -0.013,
p
< 0.001). Compared to non-operative management, laparoscopic cholecystectomy for acute cholecystitis was significantly associated with lower rates of preterm delivery, labor, or abortion (OR 0.410,
p
< 0.001). Each day that laparoscopic cholecystectomy was delayed significantly associated with an increased risk of fetal complications (OR 1.173,
p
< 0.001).
Conclusions
This nationwide study exhibits significant trends favoring surgical management of acute cholecystitis during pregnancy. Although further studies are still warranted, early laparoscopic cholecystectomy should be considered in pregnant patients with acute cholecystitis.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
ERp44 retrieves some endoplasmic reticulum (ER)-resident enzymes and immature oligomers of secretory proteins from the Golgi. Association of ERp44 with its clients is regulated by pH-dependent ...mechanisms, but the molecular details are not fully understood. Here we report high-resolution crystal structures of human ERp44 at neutral and weakly acidic pH. These structures reveal key regions in the C-terminal tail (C tail) missing in the original crystal structure, including a regulatory histidine-rich region and a subsequent extended loop. The former region forms a short α-helix (α16), generating a histidine-clustered site (His cluster). At low pH, the three Trx-like domains of ERp44 (“a,” “b,” and “b′”) undergo significant rearrangements, likely induced by protonation of His157 located at the interface between the a and b domains. The α16-helix is partially unwound and the extended loop is disordered in weakly acidic conditions, probably due to electrostatic repulsion between the protonated histidines in the His cluster. Molecular dynamics simulations indicated that helix unwinding enhances the flexibility of the C tail, disrupting its normal hydrogen-bonding pattern. The observed pH-dependent conformational changes significantly enlarge the positively charged regions around the client-binding site of ERp44 at low pH. Mutational analyses showed that ERp44 forms mixed disulfides with specific cysteines residing on negatively charged loop regions of Ero1α. We propose that the protonation states of the essential histidines regulate the ERp44–client interaction by altering the C-tail dynamics and surface electrostatic potential of ERp44.
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BFBNIB, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
The formation of a disulfide bond is a critical step in the folding of numerous secretory and membrane proteins and catalyzed in vivo. A variety of mechanisms and protein structures have evolved to ...catalyze oxidative protein folding. Those enzymes that directly interact with a folding protein to accelerate its oxidative folding are mostly thiol‐disulfide oxidoreductases that belong to the thioredoxin superfamily. The enzymes of this class often use a CXXC active‐site motif embedded in their thioredoxin‐like fold to promote formation, isomerization, and reduction of a disulfide bond in their target proteins. Over the past decade or so, an increasing number of substrates of the thiol‐disulfide oxidoreductases that are present in the ER of mammalian cells have been discovered, revealing that the enzymes play unexpectedly diverse physiological functions. However, functions of some of these enzymes still remain unclear due to the lack of information on their substrates. Here, we review the methods used by researchers to identify the substrates of these enzymes and provide data that show the importance of using trichloroacetic acid in sample preparation for the substrate identification, hoping to aid future studies. We particularly focus on successful studies that have uncovered physiological substrates and functions of the enzymes in the periplasm of Gram‐negative bacteria and the endoplasmic reticulum of mammalian cells. Similar approaches should be applicable to enzymes in other cellular compartments or in other organisms.
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FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Oxidative protein folding occurs primarily in the mammalian endoplasmic reticulum, enabled by a diverse network comprising more than 20 members of the protein disulfide isomerase (PDI) family and ...more than five PDI oxidases. Although the canonical disulfide bond formation pathway involving Ero1α and PDI has been well-studied so far, the physiological roles of the newly identified PDI oxidases, glutathione peroxidase-7 (GPx7) and -8 (GPx8), are only poorly understood. We here demonstrated that human GPx7 has much higher reactivity with H2O2 and hence greater PDI oxidation activity than human GPx8. The high reactivity of GPx7 is due to the presence of a catalytic tetrad at the redox-active site, which stabilizes the sulfenylated species generated upon the reaction with H2O2. Although it was previously postulated that GPx7 catalysis involved a highly reactive peroxidatic cysteine that can be sulfenylated by H2O2, we revealed that a resolving cysteine instead regulates the PDI oxidation activity of GPx7. We also determined that GPx7 formed complexes preferentially with PDI and P5 in H2O2-treated cells. Altogether, these results suggest that human GPx7 functions as an H2O2-dependent PDI oxidase in cells, whereas PDI oxidation may not be the central physiological role of human GPx8.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
The biological kingdoms have evolved elaborate systems that ensure the catalysis of protein disulfide bond (dsb) formation in the cell. Coexisting in the periplasm of Escherichia coli are the ...DsbA-DsbB disulfide-introducing and DsbC-DsbD disulfide-isomerizing pathways, which promote the oxidative folding of secreted proteins. Recent structural studies of DsbB have illuminated conformational dynamics involved in the effective oxidation of the extremely reduction-prone oxidase, DsbA, as well as the structure of the reaction centre involved in protein Dsb formation de novo in conjunction with ubiquinone. Extensive genetic and biochemical analysis has recently provided insight into how DsbD transports electrons from cytosolic thioredoxin to periplasmic DsbC. To a great extent, the molecular mechanisms of the Dsb enzyme system in E. coli have been elucidated, and are applicable to the study of protein disulfide formation systems in other organisms.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Fluorescent Zn2+ probes used for the quantitative analysis of labile Zn2+ concentration (Zn2+) in target organelles are crucial for understanding the role of Zn2+ in biological processes. Although ...several fluorescent Zn2+ probes have been developed to date, there is still a lack of consensus concerning the Zn2+ in intracellular organelles. In this study, we describe the development of ZnDA-1H, a small-molecule fluorescent probe for Zn2+, which exhibits less pH sensitivity, high Zn2+ selectivity, and large fluorescence enhancement upon binding to Zn2+. Through protein labeling technology, ZnDA-1H was precisely targeted in various intracellular organelles, such as the nucleus, mitochondria, endoplasmic reticulum, and Golgi apparatus. ZnDA-1H exhibited a reversible fluorescence response toward labile Zn2+ in these organelles in live cells. Using this probe, the Zn2+ in the Golgi apparatus was estimated to be 25 ± 1 nM, suggesting that labile Zn2+ plays a physiological role in the secretory pathway.
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•ZnDA-1H is a “turn-on” type green fluorescent probe with high Zn2+ selectivity•The Kd toward Zn2+ and emission of ZnDA-1H are less dependent on pH change•ZnDA-1H specifically localizes to target organelles upon conjugating with HaloTag•ZnDA-1H was applied to quantify the Zn2+ in the Golgi apparatus
Kowada et al. have developed a small-molecule fluorescent Zn2+ probe, ZnDA-1H, which can be localized specifically to intracellular organelles by utilizing a protein labeling technology. ZnDA-1H enables quantitative analysis of labile Zn2+ in the Golgi apparatus.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Introduction
Flail chest is considered a highly morbid condition with reported mortality ranging from 10 to 20%. It is often associated with other severe injuries, which may complicate management and ...interpretation of outcomes. The physiologic impact and prognosis of isolated flail chest injury is poorly defined.
Methods
This is a National Trauma Databank study. All patients from 1/2007 to 12/2014 admitted with flail chest were extracted. Patients with head or abdominal AIS ≥3, dead on arrival, or transferred, were excluded. Primary outcome was mortality; secondary outcomes were need for mechanical ventilation and pneumonia.
Results
Of the 1,047,519 patients with blunt chest injury, 14,718 (1.4%) patients presented with flail chest, and 8098 (0.77%) met inclusion criteria. The most commonly associated intrathoracic injuries were hemothorax (57.9%) and lung contusions (63.0%), while sternal fracture (8.8%) and cardiac contusion (2.5%) were less common. In total, 29.8% of patients required mechanical ventilation, and 11.2% developed pneumonia. Overall mortality was 5.6%. On multivariable analysis, age >65 and need for mechanical ventilation were independent risk factors for mortality (OR 6.02, 3.75, respectively,
p
< 0.001). Independent predictors for mechanical ventilation included cardiac or pulmonary contusion and sternal fractures (OR 3.78, 2.38, 2.29, respectively,
p
< 0.001). Need for mechanical ventilation was an independent predictor of pneumonia (OR 13.18,
p
< 0.001).
Conclusions
Mortality in isolated flail chest is much lower than previously reported. Fewer than 30% of patients require mechanical ventilation. Need for mechanical ventilation, however, is independently associated with mortality and pneumonia. Age >65 is an independent risk factor for adverse outcomes, and these patients may benefit by more aggressive monitoring and treatment.
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EMUNI, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OBVAL, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
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