Study on physisorption between phycocyanin and gold nanoparticles

Interactions between nanoparticles (NPs) and biomolecules, especially proteins, have attracted increasing attention. Photoresponsive proteins have shown high potential for optogenetic research. The combination between optogenetics and nanotechnology will bring a new biological era in which photoresponsive proteins will inevitably encounter NPs, therefore their interactions will be a key point to investigate. Here, we have systematically studied the interactions between a photoresponsive protein (called phycocyanin, PC) and a typical kind of amphiphilic polymer‐coated gold NPs (AP–AuNPs) using fluorescence quenching methods. The results showed that the binding constant between PCs and AP–AuNPs is 4.427 × 10⁶ M^–1 with a positive cooperativity, and the robust affinity was hydrophobic interaction driven mortise–tenon conjugation, which could even resist gel electrophoresis. These results could also shed light on potential designs for building up artificial protein–NP light‐harvesting systems.     

Nanostructured molecular films and nanoparticles with photoactivable functionalities

In the massively explored arena of the modern material science the creation of two- and three dimensional nanostructured molecular systems able to perform specific functions under light inputs is emerging as one of the most exciting research goals in the perspective of "intelligent" nanocomposite devices. In our laboratories we have been working on the design and fabrication of photoresponsive molecular thin films and nanoparticles over the last few years and some of the most representative examples will be illustrated in this contribution. Possible applications of these light-controlled nanosystems with particular emphasis to their impact in biological and biomedical research will be also described.     

Microfluidic technologies for studying synthetic circuits

Advances in synthetic biology have augmented the available toolkit of biomolecular modules, allowing engineering and manipulation of signaling in a variety of organisms, ranging in complexity from single bacteria and eukaryotic cells to multi-cellular systems. The richness of synthetic circuit outputs can be dramatically enhanced by sophisticated environmental control systems designed to precisely pattern spatial-temporally heterogeneous environmental stimuli controlling these circuits. Moreover, the performance of the synthetic modules and 'blocks' needed to assemble more complicated networks requires more complete characterization as a function of arbitrarily complex environmental inputs. Microfluidic technologies are poised to meet these needs through a variety of innovative designs capitalizing on the unique benefits of manipulating fluids on the micro-scales and nano-scales. This review discusses the utility of microfluidics for the study of synthetic circuits and highlights recent work in the area.     

Importance of pause between spider courtship vibrations and general problems using synthetic stimuli in behavioural studies

Effectiveness of natural stimuli (recorded and replayed male vibratory courtship signals) in eliciting a female's vibratory response was compared with that of synthetic stimuli in a plant-dwelling spider (Cupiennius salei). Specifically, the role of interseries (pauses between consecutive male series) was evaluated. Effectiveness was assessed by the number of responding females and the number of responses per female per stimulus. In case of both synthetic and natural stimuli, effectiveness increases with increasing duration of interseries in both measures. Interseries are thus represented in the female's innate releasing mechanism. However, synthetic stimuli are less effective (about 55%) than natural stimuli (set 100%). With decreasing interseries, the number of female responses per unit time sharply increases in case of playback of natural stimuli, but only slightly in case of synthetic stimuli. Since males courting on plants decrease interseries as soon as a female responds, thus facilitating their orientation, the data obtained with natural stimuli much better fit natural male courtship behaviour. This finding brings into question the validity of the synthetic stimulus. In the bioacoustics literature there is neither consensus on the effectiveness of a synthetic stimulus necessary to qualify it for behavioural analysis nor on how to measure effectiveness. We propose three measures : the percentage of responding animals, the action pattern of the response and the number of responses. It may be crucial, particularly when addressing evolutionary questions, to use either natural stimuli or synthetic stimuli (nearly) as effective as natural stimuli to avoid biased sampling of experimental animals and results difficult to interpret in biological terms.     

Systems biology as a foundation for genome-scale synthetic biology

As the ambitions of synthetic biology approach genome-scale engineering, comprehensive characterization of cellular systems is required, as well as a means to accurately model cell-scale molecular interactions. These requirements are coincident with the goals of systems biology and, thus, systems biology will become the foundation for genome-scale synthetic biology. Systems biology will form this foundation through its efforts to reconstruct and integrate cellular systems, develop the mathematics, theory and software tools for the accurate modeling of these integrated systems, and through evolutionary mechanisms. As genome-scale synthetic biology is so enabled, it will prove to be a positive feedback driver of systems biology by exposing and forcing researchers to confront those aspects of systems biology which are inadequately understood.     

Regulating gene expression with light-activated oligonucleotides

Since the development of light-responsive amino acids, the activity of numerous biomolecules has been photomodulated in biochemical, biophysical, and cellular assays. Biological problems of even greater complexity motivate the development of quantitative methods for controlling gene activity with high spatial and temporal resolution, using light as an external trigger. Photoresponsive DNA and RNA oligonucleotides would optimally serve this purpose, but have proven difficult to expand from proofs-of-concept to in vivo experiments. Until recently, the development of this technology was limited by the synthesis of oligonucleotides whose function could be significantly modulated with near-UV light. New synthetic protocols and strategies for both up- and down-regulating gene activity finally make it possible to address biological considerations. In the near future, we can expect photoresponsive DNA and RNA molecules that are relatively non-toxic, nuclease-resistant, and maintain their specificity and activity in vivo. Quantitative, laser-initiated methods for controlling DNA and RNA function will illuminate new areas in cell and developmental biology.     

合成生物学伦理、法律与社会问题探讨

合成生物学是以基因组学、系统生物学知识和分子生物学技术为基础,综合了科学与工程的一门新兴交叉学科。它使生命科学和生物技术研发进入了以人工设计、合成自然界中原本不曾出现的人造生命体系,以及对这些人工体系进行体内、体外优化,或利用这些人造生命体系研究自然生命规律为目标的新时代。然而,合成生物学研究在迅速发展、表现出巨大潜力和应用前景的同时,也引发了社会各界对相关社会、伦理、安全,以及知识产权等问题的重视与讨论。就世界各国针对合成生命对传统意义上生命概念的挑战、合成生物学产品存在的潜在风险危害、合成生物学研究的风险评估与监管等问题进行回顾综述和相关探讨。     

Engineering key components in a synthetic eukaryotic signal transduction pathway

Signal transduction underlies how living organisms detect and respond to stimuli. A goal of synthetic biology is to rewire natural signal transduction systems. Bacteria, yeast, and plants sense environmental aspects through conserved histidine kinase (HK) signal transduction systems. HK protein components are typically comprised of multiple, relatively modular, and conserved domains. Phosphate transfer between these components may exhibit considerable cross talk between the otherwise apparently linear pathways, thereby establishing networks that integrate multiple signals. We show that sequence conservation and cross talk can extend across kingdoms and can be exploited to produce a synthetic plant signal transduction system. In response to HK cross talk, heterologously expressed bacterial response regulators, PhoB and OmpR, translocate to the nucleus on HK activation. Using this discovery, combined with modification of PhoB (PhoB‐VP64), we produced a key component of a eukaryotic synthetic signal transduction pathway. In response to exogenous cytokinin, PhoB‐VP64 translocates to the nucleus, binds a synthetic PlantPho promoter, and activates gene expression. These results show that conserved‐signaling components can be used across kingdoms and adapted to produce synthetic eukaryotic signal transduction pathways.     

RNA synthetic biology

RNA molecules play important and diverse regulatory roles in the cell by virtue of their interaction with other nucleic acids, proteins and small molecules. Inspired by this natural versatility, researchers have engineered RNA molecules with new biological functions. In the last two years efforts in synthetic biology have produced novel, synthetic RNA components capable of regulating gene expression in vivo largely in bacteria and yeast, setting the stage for scalable and programmable cellular behavior. Immediate challenges for this emerging field include determining how computational and directed-evolution techniques can be implemented to increase the complexity of engineered RNA systems, as well as determining how such systems can be broadly extended to mammalian systems. Further challenges include designing RNA molecules to be sensors of intracellular and environmental stimuli, probes to explore the behavior of biological networks and components of engineered cellular control systems.     

Can we build synthetic,multicellular systems by controlling developmental signaling in space and time?

Using biological machinery to make new, functional molecules is an exciting area in chemical biology. Complex molecules containing both 'natural' and 'unnatural' components are made by processes ranging from enzymatic catalysis to the combination of molecular biology with chemical tools. Here, we discuss applying this approach to the next level of biological complexity -- building synthetic, functional biotic systems by manipulating biological machinery responsible for development of multicellular organisms. We describe recent advances enabling this approach, including first, recent developmental biology progress unraveling the pathways and molecules involved in development and pattern formation; second, emergence of microfluidic tools for delivering stimuli to a developing organism with exceptional control in space and time; third, the development of molecular and synthetic biology toolsets for redesigning or de novo engineering of signaling networks; and fourth, biological systems that are especially amendable to this approach.     

Computer-aided design of biological circuits using TinkerCell

Synthetic biology is an engineering discipline that builds on modeling practices from systems biology and wet-lab techniques from genetic engineering. As synthetic biology advances, efficient procedures will be developed that will allow a synthetic biologist to design, analyze, and build biological networks. In this idealized pipeline, computer-aided design (CAD) is a necessary component. The role of a CAD application would be to allow efficient transition from a general design to a final product. TinkerCell is a design tool for serving this purpose in synthetic biology. In TinkerCell, users build biological networks using biological parts and modules. The network can be analyzed using one of several functions provided by TinkerCell or custom programs from third-party sources. Since best practices for modeling and constructing synthetic biology networks have not yet been established, TinkerCell is designed as a flexible and extensible application that can adjust itself to changes in the field.     

The Role of the Biochemical and Biophysical Environment in Chondrogenic Stem Cell Differentiation Assays and Cartilage Tissue Engineering

The field of regenerative medicine offers hope for the development of a cell-based therapy for the repair of articular cartilage (AC). Yet, the greatest challenge in the use of stem cells for tissue repair, is understanding how the cells respond to stimuli and using that knowledge to direct cell fate. Novel methods that utilize stem cells in cartilage regeneration will require specific spatio-temporal controls of the biochemical and biophysical signaling environments. Current chondrogenic differentiation research focuses on the roles of biochemical stimuli like growth factors, hormones, and small molecules, and the role of the physical environment and mechanical stimuli, such as compression and shear stress, which likely act through mechanical receptors. Numerous signals are associated with chondrogenic-like activity of cells in different systems, however many variables for a controlled method still need to be optimized; e.g., spatial and temporal application of the stimuli, and time of transplantation of an engineered construct. Understanding the necessary microenvironmental signals for cell differentiation will advance cell therapy for cartilage repair.     

Designing de novo: interdisciplinary debates in synthetic biology

Synthetic biology is often presented as a promissory field that ambitions to produce novelty by design. The ultimate promise is the production of living systems that will perform new and desired functions in predictable ways. Nevertheless, realizing promises of novelty has not proven to be a straightforward endeavour. This paper provides an overview of, and explores the existing debates on, the possibility of designing living systems de novo as they appear in interdisciplinary talks between engineering and biological views within the field of synthetic biology. To broaden such interdisciplinary debates, we include the views from the social sciences and the humanities and we point to some fundamental sources of disagreement within the field. Different views co-exist, sometimes as controversial tensions, but sometimes also pointing to integration in the form of intermediate positions. As the field is emerging, multiple choices are possible. They will inform alternative trajectories in synthetic biology and will certainly shape its future. What direction is best is to be decided in reflexive and socially robust ways.     

Using Video Playback to Study Sexual Communication

Many sexually-selected phenotypes involve some form of visual communication. Video-playback techniques are a powerful new tool for studying visual signaling systems. Individual aspects of complex stimuli can be reliably manipulated, and stimuli can be repeatedly presented without appreciable variation in their properties. Experimenters can also construct signals which are biologically impossible, but which can be used to ask critical questions about how stimuli are perceived. Video-playback studies of sexual selection are reviewed in the context of how the methodology can be used to extend the range of questions addressed by conventional techniques. Four major issues are discussed with consideration for future studies: (1) the background against which a stimulus is presented; (2) the illumination of the stimulus; (3) the problem of pseudoreplication; and (4) experimental design considerations, including controlling for side biases and order effects and selecting appropriate response assays. Using synthetic animations may address many of these concerns.     

Engineering scalable biological systems

Synthetic biology is focused on engineering biological organisms to study natural systems and to provide new solutions for pressing medical, industrial and environmental problems. At the core of engineered organisms are synthetic biological circuits that execute the tasks of sensing inputs, processing logic and performing output functions. In the last decade, significant progress has been made in developing basic designs for a wide range of biological circuits in bacteria, yeast and mammalian systems. However, significant challenges in the construction, probing, modulation and debugging of synthetic biological systems must be addressed in order to achieve scalable higher-complexity biological circuits. Furthermore, concomitant efforts to evaluate the safety and biocontainment of engineered organisms and address public and regulatory concerns will be necessary to ensure that technological advances are translated into real-world solutions.     

合成生物学技术在环境保护中的应用

合成生物学是一个基于生物学和工程学原理的科学领域,其目的是重新设计和重组微生物,以优化或创建具有增强功能的新生物系统。该领域利用分子工具、系统生物学和遗传框架的重编程,从而构建合成途径以获得具有替代功能的微生物。传统上,合成生物学方法通常旨在开发具有成本效益的微生物细胞工厂进而从可再生资源中生产化学物质。然而,近年来合成生物学技术开始在环境保护中发挥着更直接的作用。本综述介绍了基因工程中的合成生物学工具,讨论了基于基因工程的微生物修复策略,强调了合成生物学技术可以通过响应特定污染物进行生物修复来保护环境。其中,规律间隔成簇短回文重复序列(Clustered Regularly Interspersed Short Palindromic Repeats, CRISPR)技术在基因工程细菌和古细菌的生物修复中得到了广泛应用,生物修复领域也出现了很多新的先进技术,包括生物膜工程、人工微生物群落的构建、基因驱动、酶和蛋白质工程等。有了这些新的技术和工具,生物修复将成为当今最好和最有效的污染物去除方式之一。     

Photoresponsive poly(S-(o-nitrobenzyl)-L-cysteine)-b-PEO from a L-cysteine N-carboxyanhydride monomer: synthesis, self-assembly, and phototriggered drug release

A photoresponsive S-(o-nitrobenzyl)-l-cysteine N-carboxyanhydride (NBC-NCA) monomer was for the first time designed, and the related poly(S-(o-nitrobenzyl)-l-cysteine)-b-poly(ethylene glycol) (PNBC-b-PEO) block copolymers were synthesized from the ring-opening polymerization (ROP) of NBC-NCA in DMF solution at 25 °C. Their molecular structures, physical properties, photoresponsive self-assembly, and drug release of PNBC-b-PEO were thoroughly investigated. The β-sheet conformational PNBC block within copolymers presented a thermotropic liquid crystal phase behavior, and the crystallinity of PEO block was progressively suppressed over the PNBC composition. The characteristic absorption peaks of these copolymers at about 310 and 350 nm increased over UV irradiation time and then leveled off, indicating that the o-nitrobenzyl groups were gradually photocleaved from copolymers until the completion of photocleavage. The PNBC-b-PEO copolymers self-assembled into spherical nanoparticles in aqueous solution, presenting a photoresponsive self-assembly behavior, together with a size reduction of nanoparticles after irradiation. The anticancer drug doxorubicin can be released in a controlled manner by changing the light irradiation time, which was induced by gradually photocleaving the PNBC core of nanoparticles. This work provides a facile strategy not only for the synthesis of photoresponsive polypeptide-based block copolymers but also for the fabrication of photoresponsive nanomedicine potential for anticancer therapy.     

A Spherical Model For Color Vision

An analysis of the general principles of the organization of analyzers as systems consisting of detectors shows that in such a system a stimulus may be presented as an n-dimension vector whose components are the respective excitations in selected detectors. Then the difference between two stimuli is defined by the angle between the two vectors representing them. Accordingly, the whole range of stimuli that neurons are able to discriminate may be plotted on an n-dimension sphere [4]. If we assume that for some stimuli, the difference discriminable by neurons will be directly correlated with the magnitude of the subjective difference between the stimuli, the assessment of the difference between two stimuli in a psychophysical experiment may be regarded as an arc on an n-dimension sphere of constant radius.     

The Effect of Variation in Prey Movement on the Predatory Response of Jacky Lizards (Amphibolurus muricatus)

The sensory systems of animals have evolved to meet the demands of functionally critical events. Animals that rely on visual motion cues must ignore irrelevant movement and only attend to certain characteristics that warrant further consideration. For the Australian jacky lizard ( Amphibolurus muricatus ), movement is essential for detecting potential prey. Here we examine whether differences in the actual motion characteristics of a simulated prey item influence predatory behaviour. We begin with direct observations of responses to live prey items to define an ordinal scale for subsequent video playback experiments involving a synthetic prey item (an animated cricket). In expt 1, we show that the responses of lizards to the synthetic prey were matched to those given in response to video of an actual cricket. In expt 2 we manipulated the movement patterns of the synthetic cricket based on motion analysis of actual prey movement. Manipulating motion characteristics did not influence the level of predatory behaviour observed, however, lizards showed sustained predatory behaviour to stimuli with speed characteristics that were matched to those of real crickets. We discuss the possibility that recent experience of prey movement in captivity has influenced the foraging behaviour of these lizards.