A synthetic biology approach to self-regulatory recombinant protein production in Escherichia coli

Background: Recombinant protein production is a process of great industrial interest, with products that range from pharmaceuticals to biofuels. Since high level production of recombinant protein imposes significant stress in the host organism, several methods have been developed over the years to optimize protein production. So far, these trial-and-error techniques have proved laborious and sensitive to process parameters, while there has been no attempt to address the problem by applying Synthetic Biology principles and methods, such as integration of standardized parts in novel synthetic circuits. Results: We present a novel self-regulatory protein production system that couples the control of recombinant protein production with a stress-induced, negative feedback mechanism. The synthetic circuit allows the down-regulation of recombinant protein expression through a stress-induced promoter. We used E. coli as the host organism, since it is widely used in recombinant processes. Our results show that the introduction of the self-regulatory circuit increases the soluble/insoluble ratio of recombinant protein at the expense of total protein yield. To further elucidate the dynamics of the system, we developed a computational model that is in agreement with the observed experimental data, and provides insight on the interplay between protein solubility and yield. Conclusion: Our work introduces the idea of a self-regulatory circuit for recombinant protein products, and paves the way for processes with reduced external control or monitoring needs. It demonstrates that the library of standard biological parts serves as a valuable resource for initial synthetic blocks that needs to be further refined to be successfully applied in practical problems of biotechnological significance. ^Finally, the development of a predictive model in conjunction with experimental validation facilitates a better understanding of the underlying dynamics and can be used as a guide to experimental design.(VLID)90663

Styrning i två franchiserelationer - En fallstudie om mål och krav i Reitan Servicehandel och Pölsemannen

Det finns en förhållandevis markant skillnad mellan styrningen av franchisetagare och konventionella resultatenheter. Detta eftersom det i franchiserelationerna inte finns någon målkonflikt i finansiellt hänseende. Vi har identifierat två relativt olikartade varianter av franchisestyrning; den omsorgsfulle despoten (Reitangruppen) samt den nonchalante kumpanen (Pölsemannen). Distinktionen mellan de båda typerna utgörs främst av i vilken grad tagarna kontrolleras, vilket även kan benämnas hård respektive lös styrning

Increased reliance on coronary perfusion for cardiorespiratory performance in seawater-acclimated rainbow trout

Salmonid ventricles are composed of spongy and compact myocardium, the latter being perfused via a coronary circulation. Rainbow trout (Oncorhynchus mykiss) acclimated to sea water have higher proportions of compact myocardium and display stroke volume-mediated elevations in resting cardiac output relative to freshwater-acclimated trout, probably to meet the higher metabolic needs of osmoregulatory functions. Here, we tested the hypothesis that cardiorespiratory performance of rainbow trout in sea water is more dependent on coronary perfusion by assessing the effects of coronary ligation on cardiorespiratory function in resting and exhaustively exercised trout acclimated to fresh water or sea water. While ligation only had minor effects on resting cardiorespiratory function across salinities, cardiac function after chasing to exhaustion was impaired, presumably as a consequence of atrioventricular block. Ligation reduced maximum O2 consumption rate by 33% and 17% in fish acclimated to sea water and fresh water, respectively, which caused corresponding 41% and 17% reductions in aerobic scope. This was partly explained by different effects on cardiac performance, as maximum stroke volume was only significantly impaired by ligation in sea water, resulting in 38% lower maximum cardiac output in seawater compared with 28% in fresh water. The more pronounced effect on respiratory performance in sea water was presumably also explained by lower blood O2 carrying capacity, with ligated seawater-acclimated trout having 16% and 17% lower haemoglobin concentration and haematocrit, respectively, relative to ligated freshwater trout. In conclusion, we show that the coronary circulation allows seawater-acclimated trout to maintain aerobic scope at a level comparable to that in fresh water

Relationship of Objectively-Measured Habitual Physical Activity to Chronic Inflammation and Fatigue in Middle-Aged and Older Adults

Habitual (non-exercise) physical activity (PA) declines with age, and aging-related increases in inflammation and fatigue may be important contributors to variability in PA

Performance Comparison of 112 Gb/s DMT, Nyquist PAM4 and Partial-Response PAM4 for Future 5G Ethernet-based Fronthaul Architecture

For a future 5G Ethernet-based fronthaul architecture, 100G trunk lines of a transmission distance up to 10 km over a standard single-mode fiber (SSMF) in combination with cheap gray optics to daisy chain cell site network interfaces are a promising cost- and power-efficient solution. For such a scenario, different intensity modulation and direct detect formats at a data rate of 112 Gb/s, namely Nyquist four-level pulse amplitude modulation (PAM4), discrete multitone transmission (DMT), and partial-response (PR) PAM4, are experimentally investigated, using a low-cost electroabsorption modulated laser, a 25G driver, and current state-of-the-art high-speed 84-GS/s CMOS digital-to-analog converter and analog-to-digital converter test chips. Each modulation format is optimized independently for the desired scenario, and their digital signal processing requirements are investigated. The performance of Nyquist PAM4 and PR PAM4 depends very much on the efficiency of pre- and postequalization. We show the necessity for at least 11 feedforward equalizer (FFE) taps for pre-emphasis and up to 41 FFE coefficients at the receiver side. In addition, PR PAM4 requires a maximum likelihood sequence estimation with four states to decode the signal back to a PAM4 signal. On the contrary, bit loading and power loading are crucial for DMT, and an FFT length of at least 512 is necessary. With optimized parameters, all modulation formats result in a very similar performances, demonstrating a transmission distance of up to 10 km over an SSMF with bit error rates below an FEC threshold of 4.4E-3, allowing error-free transmission

Urinary Tract Infections: Pediatric Primary Care Curriculum Podcast

Introduction: Demands on residents' time during training make it difficult for them to engage consistently with a primary care curriculum. In response to this, the emergency medicine and critical care fields have successfully utilized podcasting to the point where a recent study showed US emergency medicine residents ranked podcasts as the best use of their time for extracurricular education. Methods: We produced a 30-minute podcast on urinary tract infections from a primary care perspective, based on descriptors from Entrustable Professional Activity 4, "Manage acute common illnesses in the ambulatory setting." A moderator, a primary care pediatrician, and a pediatric nephrologist used a loose script of salient points, allowing for a natural evolution of the dialogue. The podcast was distributed to residents via email, along with a 7-question survey. Results: The survey was completed by 50 out of 84 residents. Ninety-two percent listened to all or part of the podcast, 98% found it educational, 93% enjoyed listening, and 74% felt more confident identifying and managing patients with possible urinary tract infections after listening. Ninety-six percent felt podcasts were a good alternative method for delivering this curriculum. One comment read, "This was great! It makes the information more accessible so that I can listen while working out or driving or just laying on the couch." Discussion: Based on this success, we are producing additional podcasts and will strive to keep them under 20 minutes, provide key summary points at the end, and improve ease of access by utilizing an RSS (rich site summary) feed

Impact of Diet and/or Exercise Intervention on Infrapatellar Fat Pad Morphology: Secondary Analysis from the Intensive Diet and Exercise for Arthritis (IDEA) Trial

The infrapatellar fat pad (IPFP) represents intra-articular adipose tissue that may contribute to intra-articular inflammation and pain by secretion of pro-inflammatory cytokines. Here we examined the impact of weight loss by diet and/or exercise interventions on IPFP volume

EFFECTS OF INTENSIVE DIET AND EXERCISE ON KNEE JOINT LOADS, INFLAMMATION, AND CLINICAL OUTCOMES AMONG OVERWEIGHT AND OBESE ADULTS WITH KNEE OSTEOARTHRITIS

Importance Knee osteoarthritis (OA), a common cause of chronic pain and disability, has biomechanical and inflammatory origins and is exacerbated by obesity. Objective To determine whether a ≥10% reduction in body weight induced by diet, with or without exercise, would improve mechanistic and clinical outcomes more than exercise alone. Design, Setting, and Participants Single-blind, 18-month, randomized clinical trial at Wake Forest University between July 2006 and April 2011. The diet and exercise interventions were center-based with options for the exercise groups to transition to a home-based program. Participants were 454 overweight and obese older community-dwelling adults (age ≥55 years with body mass index of 27-41) with pain and radiographic knee OA. Interventions Intensive diet-induced weight loss plus exercise, intensive diet-induced weight loss, or exercise. Main Outcomes and Measures Mechanistic primary outcomes: knee joint compressive force and plasma IL-6 levels; secondary clinical outcomes: self-reported pain (range, 0-20), function (range, 0-68), mobility, and health-related quality of life (range, 0-100). Results Three hundred ninety-nine participants (88%) completed the study. Mean weight loss for diet + exercise participants was 10.6 kg (11.4%); for the diet group, 8.9 kg (9.5%); and for the exercise group, 1.8 kg (2.0%). After 18 months, knee compressive forces were lower in diet participants (mean, 2487 N; 95% CI, 2393 to 2581) compared with exercise participants (2687 N; 95% CI, 2590 to 2784, pairwise difference [Δ]exercise vs diet = 200 N; 95% CI, 55 to 345; P = .007). Concentrations of IL-6 were lower in diet + exercise (2.7 pg/mL; 95% CI, 2.5 to 3.0) and diet participants (2.7 pg/mL; 95% CI, 2.4 to 3.0) compared with exercise participants (3.1 pg/mL; 95% CI, 2.9 to 3.4; Δexercise vs diet + exercise = 0.39 pg/mL; 95% CI, −0.03 to 0.81; P = .007; Δexercise vs diet = 0.43 pg/mL; 95% CI, 0.01 to 0.85, P = .006). The diet + exercise group had less pain (3.6; 95% CI, 3.2 to 4.1) and better function (14.1; 95% CI, 12.6 to 15.6) than both the diet group (4.8; 95% CI, 4.3 to 5.2) and exercise group (4.7; 95% CI, 4.2 to 5.1, Δexercise vs diet + exercise = 1.02; 95% CI, 0.33 to 1.71; Ppain = .004; 18.4; 95% CI, 16.9 to 19.9; Δexercise vs diet + exercise, 4.29; 95% CI, 2.07 to 6.50; Pfunction < .001). The diet + exercise group (44.7; 95% CI, 43.4 to 46.0) also had better physical health-related quality of life scores than the exercise group (41.9; 95% CI, 40.5 to 43.2; Δexercise vs diet + exercise = −2.81; 95% CI, −4.76 to −0.86; P = .005). Conclusions and Relevance Among overweight and obese adults with knee OA, after 18 months, participants in the diet + exercise and diet groups had more weight loss and greater reductions in IL-6 levels than those in the exercise group; those in the diet group had greater reductions in knee compressive force than those in the exercise group

Climate targets in European timber-producing countries conflict with goals on forest ecosystem services and biodiversity

The role of increased timber harvests in reaching climate mitigation targets for European countries will be limited if the protection of forest ecosystem services and biodiversity is to be achieved, suggests an empirical forest model driven by future scenarios to limit warming to 1.5 degrees C in 2100.The European Union (EU) set clear climate change mitigation targets to reach climate neutrality, accounting for forests and their woody biomass resources. We investigated the consequences of increased harvest demands resulting from EU climate targets. We analysed the impacts on national policy objectives for forest ecosystem services and biodiversity through empirical forest simulation and multi-objective optimization methods. We show that key European timber-producing countries - Finland, Sweden, Germany (Bavaria) - cannot fulfil the increased harvest demands linked to the ambitious 1.5 degrees C target. Potentials for harvest increase only exists in the studied region Norway. However, focusing on EU climate targets conflicts with several national policies and causes adverse effects on multiple ecosystem services and biodiversity. We argue that the role of forests and their timber resources in achieving climate targets and societal decarbonization should not be overstated. Our study provides insight for other European countries challenged by conflicting policies and supports policymakers