Exploring De Novo metabolic pathways from pyruvate to propionic acid

Industrial biotechnology provides an efficient, sustainable solution for chemical production. However, designing biochemical pathways based solely on known reactions does not exploit its full potential. Enzymes are known to accept non‐native substrates, which may allow novel, advantageous reactions. We have previously developed a computational program named Biological Network Integrated Computational Explorer (BNICE) to predict promiscuous enzyme activities and design synthetic pathways, using generalized reaction rules curated from biochemical reaction databases. Here, we use BNICE to design pathways synthesizing propionic acid from pyruvate. The currently known natural pathways produce undesirable by‐products lactic acid and succinic acid, reducing their economic viability. BNICE predicted seven pathways containing four reaction steps or less, five of which avoid these by‐products. Among the 16 biochemical reactions comprising these pathways, 44% were validated by literature references. More than 28% of these known reactions were not in the BNICE training dataset, showing that BNICE was able to predict novel enzyme substrates. Most of the pathways included the intermediate acrylic acid. As acrylic acid bioproduction has been well advanced, we focused on the critical step of reducing acrylic acid to propionic acid. We experimentally validated that Oye2p from Saccharomyces cerevisiae can catalyze this reaction at a slow turnover rate (10^?3 s^?1), which was unknown to occur with this enzyme, and is an important finding for further propionic acid metabolic engineering. These results validate BNICE as a pathway‐searching tool that can predict previously unknown promiscuous enzyme activities and show that computational methods can elucidate novel biochemical pathways for industrial applications. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:303–311, 2016     

The dynamic systems approach to control and regulation of intracellular networks

Systems theory and cell biology have enjoyed a long relationship that has received renewed interest in recent years in the context of systems biology. The term 'systems' in systems biology comes from systems theory or dynamic systems theory: systems biology is defined through the application of systems- and signal-oriented approaches for an understanding of inter- and intra-cellular dynamic processes. The aim of the present text is to review the systems and control perspective of dynamic systems. The biologist's conceptual framework for representing the variables of a biochemical reaction network, and for describing their relationships, are pathway maps. A principal goal of systems biology is to turn these static maps into dynamic models, which can provide insight into the temporal evolution of biochemical reaction networks. Towards this end, we review the case for differential equation models as a 'natural' representation of causal entailment in pathways. Block-diagrams, commonly used in the engineering sciences, are introduced and compared to pathway maps. The stimulus-response representation of a molecular system is a necessary condition for an understanding of dynamic interactions among the components that make up a pathway. Using simple examples, we show how biochemical reactions are modelled in the dynamic systems framework and visualized using block-diagrams.     

Prospects of nanoparticle–DNA binding and its implications in medical biotechnology

Bio-nanotechnology is a new interdisciplinary R&D area that integrates engineering and physical science with biology through the development of multifunctional devices and systems, focusing biology inspired processes or their applications, in particular in medical biotechnology. DNA based nanotechnology, in many ways, has been one of the most intensively studied fields in recent years that involves the use and the creation of bio-inspired materials and their technologies for highly selective biosensing, nanoarchitecture engineering and nanoelectronics. Increasing researches have been offered to a fundamental understanding how the interactions between the nanoparticles and DNA molecules could alter DNA molecular structure and its biochemical activities. This minor review describes the mechanisms of the nanoparticle–DNA binding and molecular interactions. We present recent discoveries and research progresses how the nanoparticle–DNA binding could vary DNA molecular structure, DNA detection, and gene therapy. We report a few case studies associated with the application of the nanoparticle–DNA binding devices in medical detection and biotechnology. The potential impacts of the nanoparticles via DNA binding on toxicity of the microorganisms are briefly discussed. The nanoparticle–DNA interactions and their impact on molecular and microbial functionalities have only drown attention in recent a few years. The information presented in this review can provide useful references for further studies on biomedical science and technology.     

Biotechnological advances on Penicillin G acylase: Pharmaceutical implications,unique expression mechanism and production strategies

In light of unrestricted use of first-generation penicillins, these antibiotics are now superseded by their semisynthetic counterparts for augmented antibiosis. Traditional penicillin chemistry involves the use of hazardous chemicals and harsh reaction conditions for the production of semisynthetic derivatives and, therefore, is being displaced by the biosynthetic platform using enzymatic transformations. Penicillin G acylase (PGA) is one of the most relevant and widely used biocatalysts for the industrial production of β-lactam semisynthetic antibiotics. Accordingly, considerable genetic and biochemical engineering strategies have been devoted towards PGA applications. This article provides a state-of-the-art review in recent biotechnological advances associated with PGA, particularly in the production technologies with an emphasis on using the Escherichia coli expression platform.     

代谢工程

代谢工程,也称途径工程,是基因工程一个重要分支,一般是多基因的基因工程,与细胞的基因调控、代谢调控和生化工程密切相关。讨论了代谢工程的应用,包括通过改变代谢流和代谢途径提高产量,改善生产过程,构建新的代谢途径和产生新的代谢产物等。     

The metabolomics of airway diseases,including COPD,asthma and cystic fibrosis

Abstract

Chronic obstructive pulmonary disease (COPD), asthma and cystic fibrosis (CF) are characterized by airway obstruction and an inflammatory process. Reaching early diagnosis and discrimination of subtypes of these respiratory diseases are quite a challenging task than other chronic illnesses. Metabolomics is the study of metabolic pathways and the measurement of unique biochemical molecules generated in a living system. In the last decade, metabolomics has already proved to be useful for the characterization of several pathological conditions and offers promises as a clinical tool. In this article, we review the current state of the metabolomics of COPD, asthma and CF with a focus on the different methods and instrumentation being used for the discovery of biomarkers in research and translation into clinic as diagnostic aids for the choice of patient-specific therapies.     

枯草芽孢杆菌在系统与合成生物技术中研究进展及工业应用

枯草芽孢杆菌Bacillus subtilis是微生物生理生化机理研究的模式菌株,也是工业应用生产小分子化合物、大宗化学品、工业酶、药物及保健品等生物制剂的良好底盘细胞.近些年,研究枯草芽孢杆菌的合成生物技术和代谢工程方法日新月异,为利用其作为底盘细胞生产目标产品提供了良好的工具和理论参考.文中综述了利用枯草芽孢杆菌为...     

Plant-Mediated Synthesis and Applications of Iron Nanoparticles

Nanoscale iron particles have attracted substantial interest due to their unique physical and chemical properties. Over the years, various physical and chemical methods have been developed to synthesize these nanostructures which are usually expensive and potentially harmful to human health and the environment. Synthesis of iron nanoparticles (INPs) by using plant extract is now of great interest in order to develop a novel and sustainable approach toward green chemistry. In this method the chemical compounds and organic solvents are replaced with phytochemicals and aqueous matrixes, respectively. Similar to any chemical and biochemical reaction, factors such as reaction temperature, concentration of iron precursor, concentration of leaf extract, and reaction time have critical effects on the reaction yield. This review focuses on the novel approaches used for green synthesis of INPs by using plant resources. The currently available statistics including the factors affecting the synthesis process and potential applications of the fabricated nanoparticles are discussed. Recommendations are also given for areas of future research in order to improve the production process.     

A structured approach for the engineering of biochemical network models, illustrated for signalling pathways

Quantitative models of biochemical networks (signal transduction cascades, metabolic pathways, gene regulatory circuits) are a central component of modern systems biology. Building and managing these complex models is a major challenge that can benefit from the application of formal methods adopted from theoretical computing science. Here we provide a general introduction to the field of formal modelling, which emphasizes the intuitive biochemical basis of the modelling process, but is also accessible for an audience with a background in computing science and/or model engineering. We show how signal transduction cascades can be modelled in a modular fashion, using both a qualitative approach--qualitative Petri nets, and quantitative approaches--continuous Petri nets and ordinary differential equations (ODEs). We review the major elementary building blocks of a cellular signalling model, discuss which critical design decisions have to be made during model building, and present a number of novel computational tools that can help to explore alternative modular models in an easy and intuitive manner. These tools, which are based on Petri net theory, offer convenient ways of composing hierarchical ODE models, and permit a qualitative analysis of their behaviour. We illustrate the central concepts using signal transduction as our main example. The ultimate aim is to introduce a general approach that provides the foundations for a structured formal engineering of large-scale models of biochemical networks.     

Bacterial sucrose isomerases: properties and structural studies

Due to their significant role in food industry, sucrose isomerases are good candidates for rational protein engineering. Hence, specific modifications in order to modify substrate affinity and selectivity, product specificity but also to adapt their catalytic properties to particular industrial process conditions, is interesting. Our work on the structural studies of the sucrose isomerase, MutB, which presents the first structural data available on a trehalulose synthase and the first experimental data on complexed forms of sucrose isomerases represents a significant advance in the understanding of these enzymes. In this review we give an overview of what is known on biochemical properties and structural aspects of different sucrose isomerases in particular those reported from bacteria.     

Features and technical applications of ω-transaminases

Chiral amines in enantiopure forms are important chemical building blocks, which are most well recognized in the pharmaceutical industries for imparting desirable biological activity to chemical entities. A number of synthetic strategies to produce chiral amines via biocatalytic as well as chemical transformation have been developed. Recently, ω-transaminase (ω-TA) has attracted growing attention as a promising catalyst which provides an environment-friendly access to production of chiral amines with exquisite stereoselectivity and excellent catalytic turnover. To obtain enantiopure amines using ω-TAs, either kinetic resolution of racemic amines or asymmetric amination of achiral ketones is employed. The latter is usually preferred because of twofold higher yield and no requirement of conversion of a ketone product back to racemic amine. However, the choice of a production process depends on several factors such as reaction equilibrium, substrate reactivity, enzyme inhibition, and commercial availability of substrates. This review summarizes the biochemical features of ω-TA, including reaction chemistry, substrate specificity, and active site structure, and then introduces recent advances in expanding the scope of ω-TA reaction by protein engineering and public database searching. We also address crucial factors to be considered for the development of efficient ω-TA processes.     

Towards comprehensive and quantitative proteomics for diagnosis and therapy of human disease

Given superior analytical features, MS proteomics is well suited for the basic investigation and clinical diagnosis of human disease. Modern MS enables detailed functional characterization of the pathogenic biochemical processes, as achieved by accurate and comprehensive quantification of proteins and their regulatory chemical modifications. Here, we describe how high‐accuracy MS in combination with high‐resolution chromatographic separations can be leveraged to meet these analytical requirements in a mechanism‐focused manner. We review the quantification methods capable of producing accurate measurements of protein abundance and posttranslational modification stoichiometries. We then discuss how experimental design and chromatographic resolution can be leveraged to achieve comprehensive functional characterization of biochemical processes in complex biological proteomes. Finally, we describe current approaches for quantitative analysis of a common functional protein modification: reversible phosphorylation. In all, current instrumentation and methods of high‐resolution chromatography and MS proteomics are poised for immediate translation into improved diagnostic strategies for pediatric and adult diseases.     

Status and developments of animal cell technology using suspension culture techniques

In this review the state of the art in animal cell technology, using suspension culture techniques is updated as far as the end of 1980. We have tried to discuss, on a broad basis, the current status and potential developments of both, the purely biological and the biochemical engineering aspects which may be important to improve the performance and design of animal cell technologies. The process economics could be considerably improved by the use of transformed animal cell substrates, the use of cheaper cultivation media and by methodological and engineering means. Most of these aspects are in the state of realization. Nevertheless, for a great variety of biological active substances which do not require so – or posttranslational processing, recombinant DNA-techniques (e.g. genetic engineering) are a promising alternative for the production of animal cell derived substances.     

Modeling cell migration in 3D

Cell migration is a multi-scale process that integrates signaling, mechanics and biochemical reaction kinetics. Various mathematical models accurately predict cell migration on 2-D surfaces, but are unable to capture the complexities of 3D migration. Additionally, quantitative 3D cell migration models have been few and far between. In this review we look and characterize various mathematical models available in literature to predict cell migration in 3-D matrices and analyze their strengths and possible changes to these models that could improve their predictive capabilities.     

Cell motility: the integrating role of the plasma membrane

The plasma membrane is of central importance in the motility process. It defines the boundary separating the intracellular and extracellular environments, and mediates the interactions between a motile cell and its environment. Furthermore, the membrane serves as a dynamic platform for localization of various components which actively participate in all aspects of the motility process, including force generation, adhesion, signaling, and regulation. Membrane transport between internal membranes and the plasma membrane, and in particular polarized membrane transport, facilitates continuous reorganization of the plasma membrane and is thought to be involved in maintaining polarity and recycling of essential components in some motile cell types. Beyond its biochemical composition, the mechanical characteristics of the plasma membrane and, in particular, membrane tension are of central importance in cell motility; membrane tension affects the rates of all the processes which involve membrane deformation including edge extension, endocytosis, and exocytosis. Most importantly, the mechanical characteristics of the membrane and its biochemical composition are tightly intertwined; membrane tension and local curvature are largely determined by the biochemical composition of the membrane and the biochemical reactions taking place; at the same time, curvature and tension affect the localization of components and reaction rates. This review focuses on this dynamic interplay and the feedbacks between the biochemical and biophysical characteristics of the membrane and their effects on cell movement. New insight on these will be crucial for understanding the motility process.     

Piezoelectric poly(vinylidene fluoride) microstructure and poling state in active tissue engineering

Tissue engineering strategies rely on suitable membranes and scaffolds, providing the necessary physicochemical stimuli to specific cells. This review summarizes the main results on piezoelectric polymers, in particular poly(vinylidene fluoride), for muscle and bone cell culture. Further, the relevance of polymer microstructure and surface charge on cell response is demonstrated. Together with the necessary biochemical cues, the proper design of piezoelectric polymers can open the way to novel and more reliable tissue engineering strategies for cells in which electromechanical stimuli are present in their environment.     

A miniaturized technique for assessing protein thermodynamics and function using fast determination of quantitative cysteine reactivity

Protein thermodynamic stability is a fundamental physical characteristic that determines biological function. Furthermore, alteration of thermodynamic stability by macromolecular interactions or biochemical modifications is a powerful tool for assessing the relationship between protein structure, stability, and biological function. High-throughput approaches for quantifying protein stability are beginning to emerge that enable thermodynamic measurements on small amounts of material, in short periods of time, and using readily accessible instrumentation. Here we present such a method, fast quantitative cysteine reactivity, which exploits the linkage between protein stability, sidechain protection by protein structure, and structural dynamics to characterize the thermodynamic and kinetic properties of proteins. In this approach, the reaction of a protected cysteine and thiol-reactive fluorogenic indicator is monitored over a gradient of temperatures after a short incubation time. These labeling data can be used to determine the midpoint of thermal unfolding, measure the temperature dependence of protein stability, quantify ligand-binding affinity, and, under certain conditions, estimate folding rate constants. Here, we demonstrate the fQCR method by characterizing these thermodynamic and kinetic properties for variants of Staphylococcal nuclease and E. coli ribose-binding protein engineered to contain single, protected cysteines. These straightforward, information-rich experiments are likely to find applications in protein engineering and functional genomics.     

Microbial lipases: at the interface of aqueous and non-aqueous media. A review

In recent times, biotechnological applications of microbial lipases in synthesis of many organic molecules have rapidly increased in non-aqueous media. Microbial lipases are the 'working horses' in biocatalysis and have been extensively studied when their exceptionally high stability in non-aqueous media has been discovered. Stability of lipases in organic solvents makes them commercially feasibile in the enzymatic esterification reactions. Their stability is affected by temperature, reaction medium, water concentration and by the biocatalyst's preparation. An optimization process for ester synthesis from pilot scale to industrial scale in the reaction medium is discussed. The water released during the esterification process can be controlled over a wide range and has a profound effect on the activity of the lipases. Approaches to lipase catalysis like protein engineering, directed evolution and metagenome approach were studied. This review reports the recent development in the field ofnon-aqueous microbial lipase catalysis and factors controlling the esterification/transesterification processes in organic media.     

The use of RNA interference for the metabolic engineering of plants (Review)

The metabolic engineering of plants is aimed at the realization of new biochemical reactions by transgenic cells. These reactions are determined by enzymes encoded by foreign or self-modified genes. Plants are considered to be the most interesting objects for metabolic engineering. Although they are characterized by the same pathways for the synthesis of basic biological compounds, plants differ by the astonishing diversity of their products: sugars, aromatic compounds, fatty acids, steroid compounds, and other biologically active substances. RNA interference aimed at modifying metabolic pathways is a powerful tool that allows for the obtainment of plants with new valuable properties. The present review discusses the main tendencies for research development directed toward the obtainment of transgenic plants with altered metabolism.     

运用《生物化学》歌曲，辅助《生物化学》教学

《生物化学》以生物体为对象,研究其生命的化学本质,是生命科学领域的核心课程。长期以来,由于生物化学课程知识点多、范围广和内容抽象,在一定程度上会影响学生学习的自信心,压抑其学习过程中的兴趣,致使学习的积极性不高。最近10～20年里,国外将科学（science）、技术学（technology）、工程学（engineering）及数学（mathematics）的教育与艺术学（arts）,特别是与艺术学中的音乐结合实施教学,形成一种所谓的STEAM (STEM + Arts) 策略,对STEM教育进行辅助,取得了不错的效果。基于以上情况,结合国内生物化学教学实际,笔者尝试将生物化学歌曲应用于课堂教学过程中,辅助教学。生物化学歌曲可以将抽象难懂的生物化学知识转变成悦耳动听的歌曲,在教学过程中能激发学生的学习兴趣,活跃课堂气氛,使学生在学习过程中爱上生物化学;在生物化学歌曲的创作过程中,能促进学生的思考创新,内化重点难点,使深奥的问题形象化;在学习过程中用歌声展现生物化学的魅力,让知识成为有趣的知识,让其成为有趣的学习者。本文介绍了国内生物化学歌曲发展壮大历程,结合具体实例从利用生物化学歌曲引入教学、理解生物化学内容、密切联系生活三方面评论了生物化学歌曲在辅助生物化学教学中的应用,并从歌词的改编、旋律的选择、歌曲的传唱、教学的设计等方面需要注意的问题进行讨论。