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1.
Behav Sci (Basel) ; 14(7)2024 Jul 10.
Artículo en Inglés | MEDLINE | ID: mdl-39062410

RESUMEN

Research on Chinese parenting styles using representative samples is limited, particularly during COVID-19, with most studies focusing on individual factors while neglecting regional influences. This study examines the impact of provincial income inequality, measured by the Gini coefficient, on parenting styles and how these effects vary across subgroups. Using data from the China Family Panel Studies (CFPS) 2020, encompassing 3768 children aged 7-16 years from 25 regions, we employed a multinomial logistic regression model to analyze the predictability of provincial income inequality on parenting styles identified through latent class analysis. Three parenting styles emerged during the first year of COVID-19: authoritarian (48.2%), autonomy granting (27.7%), and average-level undifferentiated (24.1%). A higher Gini coefficient related to a greater likelihood of parents adopting authoritarian or autonomy-granting parenting styles over average-level undifferentiated parenting. Subgroup analyses revealed a higher likelihood of adopting autonomy-granting and authoritarian parenting for male children compared to female. Mothers with lower education levels and parents in rural areas tend to favor authoritarian parenting in response to higher income inequality. This trend was less evident among more educated mothers and parents living in urban areas. These findings suggest that parenting styles in China are influenced by complex and region-specific factors.

2.
Sensors (Basel) ; 24(10)2024 May 12.
Artículo en Inglés | MEDLINE | ID: mdl-38793927

RESUMEN

Specialized cancer treatments have the potential to exploit glutamine dependence to increase patient survival rates. Glutamine diagnostics capable of tracking a patient's response to treatment would enable a personalized treatment dosage to optimize the tradeoff between treatment success and dangerous side effects. Current clinical glutamine testing requires sophisticated and expensive lab-based tests, which are not broadly available on a frequent, individualized basis. To address the need for a low-cost, portable glutamine diagnostic, this work engineers a cell-free glutamine biosensor to overcome assay background and signal-to-noise limitations evident in previously reported studies. The findings from this work culminate in the development of a shelf-stable, paper-based, colorimetric glutamine test with a high signal strength and a high signal-to-background ratio for dramatically improved signal resolution. While the engineered glutamine test is important progress towards improving the management of cancer and other health conditions, this work also expands the assay development field of the promising cell-free biosensing platform, which can facilitate the low-cost detection of a broad variety of target molecules with high clinical value.


Asunto(s)
Técnicas Biosensibles , Glutamina , Ingeniería Metabólica , Técnicas Biosensibles/métodos , Glutamina/metabolismo , Ingeniería Metabólica/métodos , Humanos , Ingeniería Genética/métodos , Papel , Colorimetría/métodos , Sistema Libre de Células
3.
Heliyon ; 10(9): e30009, 2024 May 15.
Artículo en Inglés | MEDLINE | ID: mdl-38737242

RESUMEN

Introduction: Dementia is marked by a steady decline or worsening in cognitive abilities, affecting memory, logic, and social competencies. While many studies suggest a potential link between the amount of sleep and dementia risk, the outcomes are not yet consistent. This research delved into the relationship between sleep length and bedtime on cognitive abilities using an extensive dataset from the China Family Panel Studies (CFPS) from 2014 to 2020. Methods: Data from 175,702 observations were collected, including cognitive function test data from 22,848 participants. Various cognitive tests were used to assess cognitive function. Restricted cubic spline (RCS) models were used for data analysis. Results: The optimal sleep duration for cognitive function was found to be 6-7 h, and the optimal bedtime was generally between 22:00-23:00. Longitudinal analysis revealed that sleep duration four years prior had a significant impact on current cognitive function. After accounting for various factors, those who slept for 7-8 h and over 8 h displayed lower cognitive function scores. Conversely, individuals sleeping less than 6 h had higher scores on the Vocabulary Test. Bedtime before 22:00 was associated with lower scores on the Vocabulary Test and Mathematical Test. Subgroup analyses based on age, gender, and urban residence showed variations in optimal sleep duration for different populations. Propensity Score Matching (PSM) analysis supported the findings. Conclusions: Maintaining a sleep duration of 6-7 h and a regular bedtime between 22:00-23:00 is important for optimizing cognitive performance.

4.
Sci Rep ; 14(1): 11611, 2024 05 21.
Artículo en Inglés | MEDLINE | ID: mdl-38773355

RESUMEN

The educational burden from extracurricular tutoring class has become a pressing social issue in China. This study used data from the China family panel studies (CFPS) in 2014, 2016, and 2018 to empirically analyze the impact of Internet usage on children's participation in extracurricular tutoring class. There are many factors that influence parents' decisions to enroll their children in extracurricular tutoring class. These factors include family income status, the level of importance parents place on their children's education, the marginal returns on educational investment, academic pressure, etc. However, in today's digitalized society, the widespread use of the internet will also become an important influencing factor in parents' decisions regarding educational investment. The study finds that, parents by using the Internet significantly increase the probability of enrolling their children in extracurricular tutoring class. Through mechanism regression analysis, it is concluded that internet usage has a positive influence on parents enrolling their children in extracurricular tutoring class by increasing the frequency of social interaction and raising parents' educational expectations for their children. Based on the empirical results, the following policy suggestions were proposed: 1. Schools should establish a more comprehensive after-school education service system to improve the engagement of students in compulsory education; 2. The government can enhance the accessibility and optimization of educational resources by increasing investment in education, improving the quality of in-school education, and optimizing the management and supervision of extracurricular tutoring class. This ensures that students can access high-quality educational services.


Asunto(s)
Uso de Internet , Padres , Estudiantes , Humanos , Niño , China , Masculino , Femenino , Uso de Internet/estadística & datos numéricos , Instituciones Académicas , Adulto , Adolescente , Internet/estadística & datos numéricos
5.
Environ Sci Pollut Res Int ; 31(23): 33414-33427, 2024 May.
Artículo en Inglés | MEDLINE | ID: mdl-38684607

RESUMEN

The mitigation of carbon dioxide (CO2) generated from household consumption, accounting for 52% of China's total greenhouse gas emissions, plays a pivotal role in China's pursuit of reaching a carbon peak by 2030. The study used three waves of nationally representative longitudinal data, energy statistics data, and input-output table to estimate household CO2 emissions (HCEs) in China at the micro-scale. The dynamic relationship between household consumption pattern transition and HCEs per capita was explored by applying maximum likelihood and structural equation modeling (ML-SEM) with panel data. The results indicate that per capita HCE level in a given year appears to be positively associated with HCE level for the same household in the previous year. A U-shaped relationship between consumption pattern transition and HCEs per capita was confirmed, as well as the reinforcement effect of income on the impacts of consumption pattern transition. The increase in consumption propensity, household income, share of wage-income, household asset values, and house space results in higher HCEs per capita. The family size and dependency ratio have a negative relationship with HCEs, whereas households that are female-oriented and more Internet-dependent tend to produce more CO2. Exploring the consumption transition of households is crucial for reducing CO2 emissions at the household level in China.


Asunto(s)
Dióxido de Carbono , Composición Familiar , China , Dióxido de Carbono/análisis , Humanos
6.
Sci Rep ; 14(1): 8042, 2024 04 05.
Artículo en Inglés | MEDLINE | ID: mdl-38580785

RESUMEN

Cell-free protein synthesis (CFPS) systems offer a versatile platform for a wide range of applications. However, the traditional methods for detecting proteins synthesized in CFPS, such as radioactive labeling, fluorescent tagging, or electrophoretic separation, may be impractical, due to environmental hazards, high costs, technical complexity, and time consuming procedures. These limitations underscore the need for new approaches that streamline the detection process, facilitating broader application of CFPS. By harnessing the reassembly capabilities of two GFP fragments-specifically, the GFP1-10 and GFP11 fragments-we have crafted a method that simplifies the detection of in vitro synthesized proteins called FAST (Fluorescent Assembly of Split-GFP for Translation Tests). FAST relies on the fusion of the small tag GFP11 to virtually any gene to be expressed in CFPS. The in vitro synthesized protein:GFP11 can be rapidly detected in solution upon interaction with an enhanced GFP1-10 fused to the Maltose Binding Protein (MBP:GFP1-10). This interaction produces a fluorescent signal detectable with standard fluorescence readers, thereby indicating successful protein synthesis. Furthermore, if required, detection can be coupled with the purification of the fluorescent complex using standardized MBP affinity chromatography. The method's versatility was demonstrated by fusing GFP11 to four distinct E. coli genes and analyzing the resulting protein synthesis in both a homemade and a commercial E. coli CFPS system. Our experiments confirmed that the FAST method offers a direct correlation between the fluorescent signal and the amount of synthesized protein:GFP11 fusion, achieving a sensitivity threshold of 8 ± 2 pmol of polypeptide, with fluorescence plateauing after 4 h. Additionally, FAST enables the investigation of translation inhibition by antibiotics in a dose-dependent manner. In conclusion, FAST is a new method that permits the rapid, efficient, and non-hazardous detection of protein synthesized within CFPS systems and, at the same time, the purification of the target protein.


Asunto(s)
Colorantes , Escherichia coli , Proteínas Fluorescentes Verdes/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Fluorescencia , Colorantes/metabolismo
7.
Front Public Health ; 12: 1348234, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-38590814

RESUMEN

China is actively encouraging households to replace traditional solid fuels with clean energy. Based on the Chinese Families Panel Survey (CFPS) data, this paper uses propensity scores matching with the difference-in-differences model to examine the impact of clean energy in the household sector on residents' health status, and whether such an energy transition promotes health equity by favoring relatively disadvantaged social groups. The results show that: (1) The use of cleaner cooking fuels can significantly improve residents' health status; (2) The older adult and women have higher health returns from the clean energy transition, demonstrating that, from the perspective of age and gender, the energy transition contributes to the promotion of health equity; (3) The clean energy transition has a lower or insignificant health impact on residents who cannot easily obtain clean energy or replace non-clean energy at an affordable price. Most of these individuals live in low-income, energy-poor, or rural households. Thus, the energy transition exacerbates health inequalities. This paper suggests that to reduce the cost of using clean energy and help address key issues in health inequality, Chinese government efforts should focus on improving the affordability, accessibility, and reliability of clean energy.


Asunto(s)
Contaminación del Aire Interior , Humanos , Femenino , Anciano , Contaminación del Aire Interior/análisis , Disparidades en el Estado de Salud , Reproducibilidad de los Resultados , China , Inequidades en Salud
8.
Methods Mol Biol ; 2762: 293-308, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-38315373

RESUMEN

Although membrane proteins are abundant in nature, their investigation is limited due to bottlenecks in heterologous overexpression and consequently restricted accessibility for downstream applications. In this chapter, we address these challenges by presenting a fast and straightforward synthesis platform based on eukaryotic cell-free protein synthesis (CFPS) and an efficient solubilization strategy using styrene-maleic acid (SMA) copolymers. We demonstrate CFPS of TWIK-1, a dimeric ion channel, based on Sf21 (Spodoptera frugiperda) insect lysate showing homooligomerization and N-glycosylation enabled by endoplasmic reticulum-derived microsomes. Furthermore, we employ SMA copolymers for protein solubilization, which preserves the native-like microsomal environment. This approach not only retains the solubilized protein's suitability for downstream applications but also maintains the oligomerization and glycosylation of TWIK-1 post-solubilization. We validate the solubilization procedure using autoradiography, particle size analysis, and biomolecular fluorescence assay and confirm the very efficient, structurally intact solubilization of cell-free synthesized TWIK-1.


Asunto(s)
Maleatos , Poliestirenos , Proteínas de la Membrana
9.
Int J Mol Sci ; 25(4)2024 Feb 07.
Artículo en Inglés | MEDLINE | ID: mdl-38396696

RESUMEN

The rise of antimicrobial resistance poses a significant global health threat, necessitating urgent efforts to identify novel antimicrobial agents. In this study, we undertook a thorough screening of soil-derived bacterial isolates to identify candidates showing antimicrobial activity against Gram-positive bacteria. A highly active antagonistic isolate was initially identified as Bacillus altitudinis ECC22, being further subjected to whole genome sequencing. A bioinformatic analysis of the B. altitudinis ECC22 genome revealed the presence of two gene clusters responsible for synthesizing two circular bacteriocins: pumilarin and a novel circular bacteriocin named altitudin A, alongside a closticin 574-like bacteriocin (CLB) structural gene. The synthesis and antimicrobial activity of the bacteriocins, pumilarin and altitudin A, were evaluated and validated using an in vitro cell-free protein synthesis (IV-CFPS) protocol coupled to a split-intein-mediated ligation procedure, as well as through their in vivo production by recombinant E. coli cells. However, the IV-CFPS of CLB showed no antimicrobial activity against the bacterial indicators tested. The purification of the bacteriocins produced by B. altitudinis ECC22, and their evaluation by MALDI-TOF MS analysis and LC-MS/MS-derived targeted proteomics identification combined with massive peptide analysis, confirmed the production and circular conformation of pumilarin and altitudin A. Both bacteriocins exhibited a spectrum of activity primarily directed against other Bacillus spp. strains. Structural three-dimensional predictions revealed that pumilarin and altitudin A may adopt a circular conformation with five- and four-α-helices, respectively.


Asunto(s)
Bacillus , Bacteriocinas , Bacteriocinas/genética , Bacteriocinas/farmacología , Antibacterianos/química , Cromatografía Liquida , Escherichia coli/metabolismo , Espectrometría de Masas en Tándem , Bacillus/metabolismo
10.
J Affect Disord ; 351: 442-448, 2024 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-38280570

RESUMEN

BACKGROUND: Employee mental health is linked with job satisfaction and productivity. However, there is a notable lack of empirical evidence regarding the adverse effects of Hazardous Work Environments (HWE) on employee mental health, indicating a significant gap in occupational health research. METHODS: Data from the China Family Panel Studies involving 9972 participants were utilized. Mental health was assessed using the Kessler 6 Psychological Distress Scale at baseline and the 8-item Center for Epidemiologic Studies Depression scale at follow-up. Associations between HWE exposure and mental health were explored cross-sectionally at baseline plus mediation analyses for the influence of sleep duration. Cox proportional hazards regression was used to assess the longitudinal implications of HWE exposure. FINDINGS: Participants exposed to HWE showed a higher risk of having serious psychological distress (OR (95%CI): 1.47 (1.29-1.69)) cross-sectionally at baseline, and this risk rose with increased number of HWE elements (1.30 (1.11-1.52) and 2.18 (1.75-2.72) for 1-2 and ≥ 3 elements, respectively). Longitudinally, HWE exposure was associated with an increased risk of depression (HR (95%CI): 1.10 (1.07-1.14)). Similarly, this risk risen with increased number of HWE elements (1.08 (1.06-1.09) and 1.20 (1.05-1.38) for 1-2 and ≥ 3 elements, respectively) and longer duration of exposure (1.01 (1.00-1.01)). Short sleep duration emerged as a significant mediator between HWE exposure and psychological distress. INTERPRETATION: This study illuminates a robust association between HWE exposure and psychological distress, especially under prolonged and multiple exposures. We advocate for rigorous mental health monitoring and interventions for workers in adverse working environments. FUNDING: CSTB2023NSCQ-BHX0042 and cffkyqdj.


Asunto(s)
Salud Mental , Lugar de Trabajo , Humanos , Lugar de Trabajo/psicología , Satisfacción en el Trabajo , Condiciones de Trabajo , China/epidemiología , Estrés Psicológico/epidemiología
11.
Heliyon ; 10(2): e24154, 2024 Jan 30.
Artículo en Inglés | MEDLINE | ID: mdl-38293341

RESUMEN

In today's digital era, households are empowered by digital servitization, which could potentially impact their ability to become entrepreneurs. Using data from the China Family Panel Studies (CFPS) and the National Bureau of Statistics of China, we construct the digital servitization index to analyze the impact of digital servitization on household entrepreneurship. The results show that the utilization of digital servitization by households significantly increases the likelihood of them engaging in entrepreneurial endeavors. This result remains robust after a series of robustness tests. We also find that digital servitization can help households identify entrepreneurial opportunities and access entrepreneurial resources by alleviating information and financial constraints, and increasing social capital. Further, the effect is more among households with rural residences, low material assets, elderly household heads, and household heads with low cognitive abilities, which means that digital servitization can promote underdog entrepreneurship. In conclusion, this paper provides micro-level evidence supporting the idea that digital servitization can foster household entrepreneurship, particularly among underprivileged groups. Ultimately, this paper highlights the potential of digitization as an essential resource to drive economic growth and help households in need become successful entrepreneurs.

12.
Front Public Health ; 11: 1303541, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-38074713

RESUMEN

The primary prerequisite for socioeconomic growth is good health, hence promoting residents' health is a vital objective of public policies. It is yet up for debate whether or not the digital economy (DE), which will be crucial to future economic growth, will eventually result in improvements in residents' health. Utilizing the China Family Panel Studies (CFPS) data in 2020, we explore how the DE affects residents' health. The findings reveal that residents' health is greatly enhanced by the DE. The eastern region sees a more dramatic improvement in residents' health as a result of the DE. Additionally, the DE can improve residents' health through the promotion of regional green development. The study's findings add to our knowledge of how the DE impacts residents' health while also offering recommendations for achieving universal health.


Asunto(s)
Desarrollo Económico , Política Pública , China
13.
BMC Psychol ; 11(1): 398, 2023 Nov 17.
Artículo en Inglés | MEDLINE | ID: mdl-37978396

RESUMEN

BACKGROUND: Although the tobacco epidemic is one of the greatest public health threats, the smoking cessation rate among Chinese adults is considerably lower. Personality information may indicate which treatments or interventions are more likely to be effective. China is the largest producer and consumer of tobacco worldwide. However, little is known about the association between smoking cessation and personality traits in China. AIM: This study aimed to examine the association between successful smoking cessation and personality traits among Chinese adults. METHODS: This cross-sectional study used data from the 2018 China Family Panel Studies. Probit regression models were employed to analyze the association between successful smoking cessation and personality traits stratified by sex. RESULTS: Lower scores for neuroticism (Coef.=-0.055, p < 0.1), lower scores for extraversion (Coef.=-0.077, p < 0.05), and higher scores for openness to experience (Coef.=0.045, p < 0.1) predicted being a successful male quitter after adjusting for demographics. Moreover, lower scores for conscientiousness (Coef.=-0.150, p < 0.1) predicted being a successful female quitter after adjusting for demographics. CONCLUSION: The empirical findings suggested that among Chinese men, lower levels of neuroticism, lower levels of extraversion, and higher levels of openness to experience were associated with a higher likelihood of smoking cessation. Moreover, lower levels of conscientiousness were associated with successful smoking cessation among Chinese women. These results showed that personality information should be included in smoking cessation interventions.


Asunto(s)
Cese del Hábito de Fumar , Humanos , Masculino , Adulto , Femenino , Estudios Transversales , Pueblos del Este de Asia , Personalidad , Neuroticismo , Inventario de Personalidad
14.
Nat Sci Sleep ; 15: 915-924, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-37954027

RESUMEN

Objective: The study aims to reveal the association between family context and sleep trajectories in middle-aged and elderly Chinese adults. Methods: Subjects (n=7777) aged between 40 and 65 years were selected from the China Family Panel Studies (CFPS). Latent class analysis and the multi-trajectory method were used to identify the family context and sleep trajectories from 2010 to 2018. Multinomial (polytomous) logistic regression was performed to explore the relationship between family context and sleep trajectories. Results: Five family context classes were identified according to family demographic characteristics. Simultaneously, four sleep trajectories were determined based on three sleep-related indexes. Subjects from family that had only sons or multiple-child are liable to shorten or prolong sleep duration and increase midday nap ratios compare with subjects who from family that had one or more daughters, and in future public health prevention and control, more attention could be paid to such families. Conclusion: The study found that family context is associated with sleep trajectories among middle and old Chinese adults. Subjects from families with only girls seemed to have more stable sleep trajectories, while those with one or more boys' families had unstable sleep trajectories. Further interventions would be carried out for sleep disorders, it is necessary to pay more attention to the family context, especially the number and gender of children.

15.
Int J Mol Sci ; 24(19)2023 Oct 03.
Artículo en Inglés | MEDLINE | ID: mdl-37834312

RESUMEN

Integral membrane proteins are important components of a cell. Their structural and functional studies require production of milligram amounts of proteins, which nowadays is not a routine process. Cell-free protein synthesis is a prospective approach to resolve this task. However, there are few known membrane mimetics that can be used to synthesize active membrane proteins in high amounts. Here, we present the application of commercially available "Facade" detergents for the production of active rhodopsin. We show that the yield of active protein in lipid bicelles containing Facade-EM, Facade-TEM, and Facade-EPC is several times higher than in the case of conventional bicelles with CHAPS and DHPC and is comparable to the yield in the presence of lipid-protein nanodiscs. Moreover, the effects of the lipid-to-detergent ratio, concentration of detergent in the feeding mixture, and lipid composition of the bicelles on the total, soluble, and active protein yields are discussed. We show that Facade-based bicelles represent a prospective membrane mimetic, available for the production of membrane proteins in a cell-free system.


Asunto(s)
Membrana Dobles de Lípidos , Proteínas de la Membrana , Proteínas de la Membrana/química , Membrana Dobles de Lípidos/química , Detergentes/química , Sistema Libre de Células , Micelas
16.
Biotechnol Appl Biochem ; 70(6): 2136-2149, 2023 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-37735977

RESUMEN

Cell-free protein synthesis (CFPS) system is an innovative technology with a wide range of potential applications that could challenge current thinking and provide solutions to environmental and health issues. CFPS system has been demonstrated to be a successful way of producing biomolecules in a variety of applications, including the biomedical industry. Although there are still obstacles to overcome, its ease of use, versatility, and capacity for integration with other technologies open the door for it to continue serving as a vital instrument in synthetic biology research and industry. In this review, we mainly focus on the cell-free based platform for various product productions. Moreover, the challenges in the bio-therapeutic aspect using cell-free systems and their future prospective for the improvement and sustainability of the cell free systems.


Asunto(s)
Biotecnología , Biosíntesis de Proteínas , Sistema Libre de Células , Biotecnología/métodos , Biología Sintética , Industrias
17.
BMC Public Health ; 23(1): 1611, 2023 08 24.
Artículo en Inglés | MEDLINE | ID: mdl-37612596

RESUMEN

BACKGROUND: The debate on the relationship between social capital and health is still ongoing. To enhance previous research, this study uses data drawn from China to analyse the situations in which social capital is related to good health and the various configurations that result in good health outcomes. METHODS: Using the data of China Family Panel Studies, the conditions of age, gender, marriage, education, income, structural social capital and cognitive social capital were included to analyse the sufficient and necessary conditions for achieving good general health and their different configurations using the fsQCA method. RESULTS: None of the listed conditions were prerequisites for excellent general health in terms of either their presence or their absence. The sufficiency analysis found 11 configurations with an average of 3-4 conditions per configuration; in no configuration was the condition of social capital present alone. Structured social capital and cognitive social capital exhibited negative states in configurations 1 and 2, respectively. The most prevalent factor in all configurations was the condition of age. CONCLUSIONS: The relationship between social capital and health is both positive and negative, with cognitive social capital playing a larger role in the positive relationship than structural social capital. Social capital is neither a necessary nor a sufficient condition for health, and it must be combined with a variety of other factors to promote health. A variety of methods can be used to promote an individual's health, as different populations require different approaches to good general health, and no single pathway applies to all populations. In the Chinese population, an individual's age is a significant determinant of their health status.


Asunto(s)
Salud , Capital Social , Determinantes Sociales de la Salud , Humanos , Pueblo Asiatico , China/epidemiología , Escolaridad , Promoción de la Salud
18.
Biotechnol Prog ; 39(6): e3373, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-37408088

RESUMEN

Cell-free protein synthesis (CFPS) systems are an attractive method to complement the usual cell-based synthesis of proteins, especially for screening approaches. The literature describes a wide variety of CFPS systems, but their performance is difficult to compare since the reaction components are often used at different concentrations. Therefore, we have developed a calculation tool based on amino acid balancing to evaluate the performance of CFPS by determining the fractional yield as the ratio between theoretically achievable and experimentally achieved protein molar concentration. This tool was applied to a series of experiments from our lab and to various systems described in the literature to identify systems that synthesize proteins very efficiently and those that still have potential for higher yields. The well-established Escherichia coli system showed a high efficiency in the utilization of amino acids, but interestingly, less considered systems, such as those based on Vibrio natriegens or Leishmania tarentolae, also showed exceptional fractional yields of over 70% and 90%, respectively, implying very efficient conversions of amino acids. The methods and tools described here can quickly identify when a system has reached its maximum or has limitations. We believe that this approach will facilitate the evaluation and optimization of existing CFPS systems and provides the basis for the systematic development of new CFPS systems.


Asunto(s)
Aminoácidos , Biosíntesis de Proteínas , Aminoácidos/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas/metabolismo , Sistema Libre de Células/metabolismo
19.
ACS Synth Biol ; 12(9): 2561-2577, 2023 09 15.
Artículo en Inglés | MEDLINE | ID: mdl-37490644

RESUMEN

The fragility of biological systems during storage, transport, and utilization necessitates reliable cold-chain infrastructure and limits the potential of biotechnological applications. In order to unlock the broad applications of existing and emerging biological technologies, we report the development of a novel solid-state storage platform for complex biologics. The resulting solid-state biologics (SSB) platform meets four key requirements: facile rehydration of solid materials, activation of biochemical activity, ability to support complex downstream applications and functionalities, and compatibility for deployment in a variety of reaction formats and environments. As a model system of biochemical complexity, we utilized crudeEscherichia colicell extracts that retain active cellular metabolism and support robust levels of in vitro transcription and translation. We demonstrate broad versatility and utility of SSB through proof-of-concepts for on-demand in vitro biomanufacturing of proteins at a milliliter scale, the activation of downstream CRISPR activity, as well as deployment on paper-based devices. SSBs unlock a breadth of applications in biomanufacturing, discovery, diagnostics, and education in resource-limited environments on Earth and in space.


Asunto(s)
Productos Biológicos , Proteínas , Biotecnología , Sistema Libre de Células
20.
Int J Biol Macromol ; 242(Pt 3): 125075, 2023 Jul 01.
Artículo en Inglés | MEDLINE | ID: mdl-37230450

RESUMEN

Biocatalysis can improve current bioprocesses by identifying or improving enzymes that withstand harsh and unnatural operating conditions. Immobilized Biocatalyst Engineering (IBE) is a novel strategy integrating protein engineering and enzyme immobilization as a single workflow. Using IBE, it is possible to obtain immobilized biocatalysts whose soluble performance would not be selected. In this work, Bacillus subtilis lipase A (BSLA) variants obtained through IBE were characterized as soluble and immobilized biocatalysts, and how the interactions with the support affect their structure and catalytic performance were analyzed using intrinsic protein fluorescence. Variant P5G3 (Asn89Asp, Gln121Arg) showed a 2.6-fold increased residual activity after incubation at 76 °C compared to immobilized wild-type (wt) BSLA. On the other hand, variant P6C2 (Val149Ile) showed 4.4 times higher activity after incubation in 75 % isopropyl alcohol (36 °C) compared to Wt_BSLA. Furthermore, we studied the advancement of the IBE platform by performing synthesis and immobilizing the BSLA variants using a cell-free protein synthesis (CFPS) approach. The observed differences in immobilization performance, high temperature, and solvent resistance between the in vivo-produced variants and Wt_BSLA were confirmed for the in vitro synthesized enzymes. These results open the door for designing strategies integrating IBE and CFPS to generate and screen improved immobilized enzymes from genetic diversity libraries. Furthermore, it was confirmed that IBE is a platform that can be used to obtain improved biocatalysts, especially those with an unremarkable performance as soluble biocatalysts, which wouldn't be selected for immobilization and further development for specific applications.


Asunto(s)
Enzimas Inmovilizadas , Ingeniería de Proteínas , Biocatálisis , Enzimas Inmovilizadas/química , Ingeniería de Proteínas/métodos , Lipasa/química , Solventes/química
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