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Heenam Stanley Kim - One of the best experts on this subject based on the ideXlab platform.
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High adaptability of the Omega Loop underlies the substrate-spectrum-extension evolution of a class A β-lactamase, PenL
Scientific Reports, 2016Co-Authors: Jin Myung Choi, Junghyun Hwang, Fabio Prati, Thinh Phat Cao, Sung Haeng Lee, Heenam Stanley KimAbstract:The Omega Loop in β-lactamases plays a pivotal role in substrate recognition and catalysis, and some mutations in this Loop affect the adaptability of the enzymes to new antibiotics. Various mutations, including substitutions, deletions, and intragenic duplications resulting in tandem repeats (TRs), have been associated with β-lactamase substrate spectrum extension. TRs are unique among the mutations as they cause severe structural perturbations in the enzymes. We explored the process by which TRs are accommodated in order to test the adaptability of the Omega Loop. Structures of the mutant enzymes showed that the extra amino acid residues in the Omega Loop were freed outward from the enzyme, thereby maintaining the overall enzyme integrity. This structural adjustment was accompanied by disruptions of the internal α-helix and hydrogen bonds that originally maintained the conformation of the Omega Loop and the active site. Consequently, the mutant enzymes had a relaxed binding cavity, allowing for access of new substrates, which regrouped upon substrate binding in an induced-fit manner for subsequent hydrolytic reactions. Together, the data demonstrate that the design of the binding cavity, including the Omega Loop with its enormous adaptive capacity, is the foundation of the continuous evolution of β-lactamases against new drugs.
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Deletion mutations conferring substrate spectrum extension in the class A β-lactamase
Antimicrobial Agents and Chemotherapy, 2014Co-Authors: Junghyun Hwang, Kwang-hwi Cho, Han Song, Heenam Stanley KimAbstract:We describe four new deletion mutations in a class A β-lactamase PenA in Burkholderia thailandensis, each conferring an extended substrate spectrum. Single-amino-acid deletions T171del, I173del, and P174del and a two-amino-acid deletion, R165_T167delinsP, occurred in the Omega Loop, increasing the flexibility of the binding cavity. This rare collection of mutations has significance, allowing exploration of the diverse evolutionary trajectories of β-lactamases and as potential future mutations conferring high-level ceftazidime resistance on isolates from clinical settings, compared with amino acid substitution mutations.
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Substrate Spectrum Extension of PenA in Burkholderia thailandensis with a Single Amino Acid Deletion, Glu168del
Antimicrobial Agents and Chemotherapy, 2012Co-Authors: Karan Kim, Kwang-hwi Cho, Oksung Jung, Heenam Stanley KimAbstract:We describe a deletion mutation in a class A β-lactamase, PenA, of Burkholderia thailandensis that extended the substrate spectrum of the enzyme to include ceftazidime. Glu168del was located in a functional domain called the Omega Loop causing expansion of the space in the Loop, which in turn increased flexibility at the active site. This deletion mutation represents a rare but significant alternative mechanical path to substrate spectrum extension in PenA besides more common substitution mutations.
Timothy Palzkill - One of the best experts on this subject based on the ideXlab platform.
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Systematic mutagenesis of the active site Omega Loop of TEM-1 beta-lactamase.
Journal of bacteriology, 1996Co-Authors: Joseph F. Petrosino, Timothy PalzkillAbstract:Beta-Lactamase is a bacterial protein that provides resistance against beta-lactam antibiotics. TEM-1 beta-lactamase is the most prevalent plasmid-mediated beta-lactamase in gram-negative bacteria. Normally, this enzyme has high levels of hydrolytic activity for penicillins, but mutant beta-lactamases have evolved with activity toward a variety of beta-lactam antibiotics. It has been shown that active site substitutions are responsible for changes in the substrate specificity. Since mutant beta-lactamases pose a serious threat to antimicrobial therapy, the mechanisms by which mutations can alter the substrate specificity of TEM-1 beta-lactamase are of interest. Previously, screens of random libraries encompassing 31 of 55 active site amino acid positions enabled the identification of the residues responsible for maintaining the substrate specificity of TEM-1 beta-lactamase. In addition to substitutions found in clinical isolates, many other specificity-altering mutations were also identified. Interestingly, many nonspecific substitutions in the N-terminal half of the active site Omega Loop were found to increase ceftazidime hydrolytic activity and decrease ampicillin hydrolytic activity. To complete the active sight study, eight additional random libraries were constructed and screened for specificity-altering mutations. All additional substitutions found to alter the substrate specificity were located in the C-terminal half of the active site Loop. These mutants, much like the N-terminal Omega Loop mutants, appear to be less stable than the wild-type enzyme. Further analysis of a 165-YYG-167 triple mutant, selected for high levels of ceftazidime hydrolytic activity, provides an example of the correlation which exists between enzyme instability and increased ceftazidime hydrolytic activity in the ceftazidime-selected Omega Loop mutants.
Maria Luger - One of the best experts on this subject based on the ideXlab platform.
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Vitamin D3 Loading Is Superior to Conventional Supplementation After Weight Loss Surgery in Vitamin D-Deficient Morbidly Obese Patients: a Double-Blind Randomized Placebo-Controlled Trial
Obesity Surgery, 2016Co-Authors: Maria Luger, Renate Kruschitz, Felix B. Langer, Gerhard Prager, Friedrich Hoppichler, Karin Schindler, Christian Kienbacher, Enikö Kállay, Stefan Traussnigg, Michael TraunerAbstract:Background Bariatric patients often suffer from vitamin D deficiency (VDD), and both, morbid obesity and VDD, are related to non-alcoholic fatty liver disease. However, limited data are available regarding best strategies for treating VDD, particularly, in bariatric patients undergoing Omega-Loop gastric bypass (OLGB). Therefore, we examined the efficacy and safety of a forced vitamin D dosing regimen and intervention effects in liver fibrotic patients.
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The Effect of Roux-en-Y vs. Omega-Loop Gastric Bypass on Liver, Metabolic Parameters, and Weight Loss.
Obesity Surgery, 2016Co-Authors: Renate Kruschitz, Maria Luger, Felix B. Langer, Gerhard Prager, Karin Schindler, Christian Kienbacher, Michael Trauner, Carmen Klammer, Michael Krebs, Bernhard LudvikAbstract:Background Omega-Loop gastric bypass (OLGB) results in weight loss (WL) but data on its impact on liver and glucose metabolism compared to Roux-en-Y gastric bypass (RYGB) is lacking. Therefore, the aim of this study was to compare the development of hepatic and metabolic markers as well as WL between the above-mentioned surgical groups during the first postoperative year.
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The link between obesity and vitamin D in bariatric patients with Omega-Loop gastric bypass surgery - a vitamin D supplementation trial to compare the efficacy of postoperative cholecalciferol loading (LOAD): study protocol for a randomized controlle
Trials, 2015Co-Authors: Maria Luger, Renate Kruschitz, Rodrig Marculescu, Friedrich Hoppichler, Christian Kienbacher, Carmen Klammer, Helmuth Haslacher, Enikö Kállay, Melanie Kral, Felix B. LangerAbstract:Beyond its classical role in calcium homoeostasis and bone metabolism, vitamin D deficiency has been found to be associated with several diseases, including diabetes, non-alcoholic fatty liver disease, and even obesity itself. Importantly, there are limited data on therapeutic strategies for vitamin D deficiency in bariatric patients, and the procedure-specific guidelines may not be sufficient. To improve long-term outcomes, nutritional screening and appropriate supplementation to prevent nutrient deficiencies are urgently needed. Therefore, the aim of this study is to examine effects and safety of a forced dosing regimen of vitamin D versus conventional dose supplementation on vitamin D levels and other parameters in bariatric patients. The study includes loading plus repeat dosing compared with repeated administration of vitamin D without a loading dose, according to guidelines, in a prospective, double-blind, randomized controlled trial. Up to a triple oral loading dose is given on day 1, then 2 and 4 weeks after surgery (100,000 IU dose each time), followed by an oral maintenance dose (3420 IU/day). The control group (n = 25) will receive placebo, followed by administration of a standard dose (3420 IU/day). We hypothesize that a significant increase in vitamin D levels will occur in patients in the treatment group (n = 25) by 24 weeks after surgery. Further measurements are aimed at evaluating changes in inflammation, bone turnover, insulin resistance, blood pressure, liver, mental health, and gut microbiota of patients undergoing Omega-Loop gastric bypass surgery. Furthermore, possible associations between concentrations of vitamin D, the involved enzymes, or vitamin D receptor in adipose and/or liver tissues will be determined. To our knowledge, this trial is the first of its kind with this type of vitamin D supplementation in bariatric patients. Its major strength is the design and implementation of evaluation of influencing factors such as liver function, bone health, inflammation, insulin resistance, blood pressure, symptoms of depression, or microbiota. This alternative vitamin D dosing regimen has the potential to be a safe, fast, evidence-based treatment of vitamin D deficiency in bariatric patients. Owing to the increasing number of bariatric patients, it is also of interest to elucidate the link between obesity and vitamin D. ClinicalTrials.gov identifier: NCT02092376 . Registered on 17 March 2014.
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Effects of Omega-Loop Gastric Bypass on Vitamin D and Bone Metabolism in Morbidly Obese Bariatric Patients
Obesity Surgery, 2014Co-Authors: Maria Luger, Renate Kruschitz, Felix B. Langer, Gerhard Prager, Melanie Walker, Rodrig Marculescu, Friedrich Hoppichler, Karin Schindler, Bernhard LudvikAbstract:Background Bariatric patients often suffer from nutrient deficiencies. Little is known about vitamin D levels and bone metabolism in patients undergoing Omega-Loop gastric bypass (OLGB). We, therefore, evaluated parameters of vitamin D metabolism preoperatively and during the first postoperative year.
Jean-luc Faucheron - One of the best experts on this subject based on the ideXlab platform.
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Totally Robotic Reversal of Omega-Loop Gastric Bypass to Normal Anatomy
Obesity Surgery, 2016Co-Authors: Fabian Reche, Adrian Mancini, Anne-laure Borel, Jean-luc FaucheronAbstract:Gastric bypass procedures can potentially lead to middle and long-term complications (Podnos et al. Arch Surg 138(9):957-61, 2003). For several years, Roux-en-Y gastric bypass reversal procedures performed by laparotomy or laparoscopic way have been described in literature (Moon et al. Surg Obes Relat Dis 11(4):821-6, 2015). Major complications are anastomotic ulcers, anastomotic complications or functional disorder such as dumping syndrome, hypocalcemia, severe hypoglycemia, and malnutrition (Moon et al. Surg Obes Relat Dis 11(4):821-6, 2015; Campos et al. Surg Obes Relat Dis 10(1):36-43, 2014). One-anastomosis gastric bypass (OAGB) also called Omega-Loop gastric bypass (OLGB) or mini-gastric bypass (MGB) is a technique that has demonstrated similar results to traditional Roux-en-Y procedures in terms of weight loss and postoperative quality of life (Lee et al. Ann Surg 242(1):20-8, 2005). However, in a recent description of 1000 patients, the percentage of malnutrition was 0.2 % (two patients) with an indication to revert Omega-Loop gastric bypass back into normal anatomy (Chevallier et al. Obes Surg 25(6):951-8, 2015), but technical details have not been exposed yet. The first robotic gastric bypass was published by Horgan and Vanuno in 2001 (Horgan and Vanuno J Laparoendosc Adv Surg Tech A 11(6):415-9, 2001). The present work describes for the first time a robotic procedure to reverse OLGB into normal anatomy. METHODS: We present the video report of a 69-year-old woman suffering of severe malnutrition (weight of 42 kg, body mass index of 15.8 kg/m(2), albumin 21 g/l) who had undergone laparoscopic Omega-Loop gastric bypass 2 years ago (initial weight of 104 kg and initial body mass index of 39.6 kg/m(2)). She was referred to our Bariatric Surgery Unit, and after a period of parenteral nutrition support to improve nutritional status (albumin 32 g/l), we decided in a multidisciplinary staff to perform a reversal Omega-Loop gastric bypass back into normal anatomy using the DaVinci Si™ system by Intuitive Surgical Inc ®, Sunnyvale, CA. RESULTS: In this high definition video, we present step-by-step robotic reversal of the Omega-Loop gastric bypass. The procedure began with a careful adhesiolysis of the left lobe of the liver, small gastric pouch, and Omega-Loop. Then, the gastro-jejunostomy was transected with a 45-mm Endo GIA endocutter with purple staples. The key-point was the creation of a gastro-gastric anastomosis between the small gastric pouch and the excluded stomach. Omega-Loop jejunum was resected and the anastomosis was performed in order to avoid intestinal stenosis. The operative time was 232 min. Postoperative course was uneventful and the patient was discharged in postoperative day 8. One month after the procedure, she has gained 10 kg (albumin 34 g/l) and stabilized her nutritional status without further nutritional support. CONCLUSIONS: This is the first case described in the literature of a reversal Omega-Loop gastric bypass into normal anatomy and the first description of the use of a robotic approach. This intervention is challenging, but a feasible procedure. This technology may increase the number of surgeons who are able to provide the benefits of minimal invasive surgery to their patients without the increased risks of complications associated with initial learning curves. The three-dimensional robotic vision, a stable camera, and the multiples degrees of freedom of the robotic instruments are the features that seem to provide greater surgical precision for these complex laparoscopic operations.
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Totally Robotic Reversal of Omega-Loop Gastric Bypass to Normal Anatomy
Obesity Surgery, 2016Co-Authors: Fabian Reche, Adrian Mancini, Anne-laure Borel, Jean-luc FaucheronAbstract:Background Gastric bypass procedures can potentially lead to middle and long-term complications (Podnos et al. Arch Surg 138(9):957–61, 2003). For several years, Roux-en-Y gastric bypass reversal procedures performed by laparotomy or laparoscopic way have been described in literature (Moon et al. Surg Obes Relat Dis 11(4):821–6, 2015). Major complications are anastomotic ulcers, anastomotic complications or functional disorder such as dumping syndrome, hypocalcemia, severe hypoglycemia, and malnutrition (Moon et al. Surg Obes Relat Dis 11(4):821–6, 2015; Campos et al. Surg Obes Relat Dis 10(1):36–43, 2014). One-anastomosis gastric bypass (OAGB) also called Omega-Loop gastric bypass (OLGB) or mini-gastric bypass (MGB) is a technique that has demonstrated similar results to traditional Roux-en-Y procedures in terms of weight loss and postoperative quality of life (Lee et al. Ann Surg 242(1):20–8, 2005). However, in a recent description of 1000 patients, the percentage of malnutrition was 0.2 % (two patients) with an indication to revert Omega-Loop gastric bypass back into normal anatomy (Chevallier et al. Obes Surg 25(6):951–8, 2015), but technical details have not been exposed yet. The first robotic gastric bypass was published by Horgan and Vanuno in 2001 (Horgan and Vanuno J Laparoendosc Adv Surg Tech A 11(6):415–9, 2001). The present work describes for the first time a robotic procedure to reverse OLGB into normal anatomy.
Andrew Coulson - One of the best experts on this subject based on the ideXlab platform.
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Structural basis for the inactivation of the P54 mutant of beta-lactamase from Staphylococcus aureus PC1.
Biochemistry, 1991Co-Authors: Osnat Herzberg, Geeta Kapadia, Bernardo Blanco, Tom S. Smith, Andrew CoulsonAbstract:The crystal structure of a mutant protein of a class A beta-lactamase from Staphylococcus aureus PC1, in which Asp179 is replaced by an asparagine (P54), has been determined and refined at 2.3-A resolution (1 A = 0.1 nm). The resulting crystallographic R factor [formula: see text] are the observed and calculated structure factor amplitudes) is 0.181 for 12289 reflections with I greater than or equal to sigma (I) within the 6.0-2.3-A resolution range. The mutated residue is located at the C-terminus of an extensive Loop (the Omega-Loop), remote from the active site, and results in a drastically reduced activity. Examination of the native and P54 structures reveals that the overall fold is similar, except that there is substantial disorder of the Omega-Loop of P54. This is a consequence of the elimination of a salt bridge between Asp179 and Arg164 that links the two ends of the Omega-Loop in native beta-lactamase. It is associated with a difference in side-chain conformation between Asn179 in P54 and Asp179 in the native structure. An alternate interaction occurs in P54 between Asn179 and Ala69, adjacent to the catalytic Ser70. This disorder affects catalysis since some of the disordered residues, in particular Glu166, form part of the active site.(ABSTRACT TRUNCATED AT 250 WORDS)
