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Since its first appearance in the 1960s, solid support-free liquid-liquid chromatography has played an ever-growing role in the field of natural products research. The use of the two phases of a liquid biphasic system, the mobile and stationary phases, renders the technique highly versatile and adaptable to a wide spectrum of target molecules, from hydrophobic to highly polar small molecules to proteins. Generally considered a niche technique used only for small-scale preparative separations, liquid-liquid chromatography currently lags far behind conventional liquid-solid chromatography and liquid-liquid extraction in process modeling and industrial acceptance. This review aims to expose a broader audience to this high-potential separation technique by presenting the wide variety of available operating modes and solvent systems as well as structured, model-based design approaches. Topics currently offering opportunities for further investigation are also addressed. Expected final online publication date for the Annual Review of Chemical and Biomolecular Engineering, Volume 12 is June 2021. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

Chromatin adopts different configurations that are regulated by reversible covalent modifications, referred to as epigenetic marks. Epigenetic inhibitors have been approved for clinical use to restore epigenetic aberrations that result in silencing of tumor-suppressor genes, oncogene addictions, and enhancement of immune responses. However, these drugs suffer from major limitations, such as a lack of locus selectivity and potential toxicities. Technological advances have opened a new era of precision molecular medicine to reprogram cellular physiology. The locus-specificity of CRISPR/dCas9/12a to manipulate the epigenome is rapidly becoming a highly promising strategy for personalized medicine. This review focuses on new state-of-the-art epigenome editing approaches to modify the epigenome of neoplasms and other disease models towards a more ‘normal-like state’, having characteristics of normal tissue counterparts. We highlight biomolecular engineering methodologies to assemble, regulate, and deliver multiple epigenetic effectors that maximize the longevity of the therapeutic effect, and we discuss limitations of the platforms such as targeting efficiency and intracellular delivery for future clinical applications.

© 2020 Wiley Periodicals LLC Today's Biochemical Engineer may contribute to advances in a wide range of technical areas. The recent Biochemical and Molecular Engineering XXI conference focused on “The Next Generation of Biochemical and Molecular Engineering: The role of emerging technologies in tomorrow's products and processes”. On the basis of topical discussions at this conference, this perspective synthesizes one vision on where investment in research areas is needed for biotechnology to continue contributing to some of the world's grand challenges.

The increased availability of user-friendly and accessible computational tools for biomolecular modeling would expand the reach and application of biomolecular engineering and design. For protein modeling, one key challenge is to reduce the complexities of 3D protein folds to sets of parametric equations that nonetheless capture the salient features of these structures accurately. At present, this is possible for a subset of proteins, namely, repeat proteins. The α-helical coiled coil provides one such example, which represents ≈ 3-5% of all known protein-encoding regions of DNA. Coiled coils are bundles of α helices that can be described by a small set of structural parameters. Here we describe how this parametric description can be implemented in an easy-to-use web application, called CCBuilder 2.0, for modeling and optimizing both α-helical coiled coils and polyproline-based collagen triple helices. This has many applications from providing models to aid molecular replacement for X-ray crystallography, in silico model building and engineering of natural and designed protein assemblies, and through to the creation of completely de novo "dark matter" protein structures. CCBuilder 2.0 is available as a web-based application, the code for which is open-source and can be downloaded freely. http://coiledcoils.chm.bris.ac.uk/ccbuilder2. Lay summary We have created CCBuilder 2.0, an easy to use web-based application that can model structures for a whole class of proteins, the α-helical coiled coil, which is estimated to account for 3-5% of all proteins in nature. CCBuilder 2.0 will be of use to a large number of protein scientists engaged in fundamental studies, such as protein structure determination, through to more-applied research including designing and engineering novel proteins that have potential applications in biotechnology.

Molecular hydrogen is a promising currency in the future energy economy due to the uncertain availability of finite fossil fuel resources and environmental effects from their combustion. It also has important uses in the production of fertilizers and platform chemicals as well as in upgrading conventional fuels. Conventional methods for producing molecular hydrogen from natural gas produce carbon dioxide and use a finite resource as feedstock. However, these issues can be overcome by using light energy from the Sun combined with microorganisms and their molecular machinery capable of photosynthesis. In the presence of light, the proteins involved in photosynthesis coupled with appropriate catalysts in higher plants, algae, and cyanobacteria can produce molecular hydrogen, and optimization via genetic modifications and biomolecular engineering further improves production rates. In this review, we will discuss techniques that have been utilized to improve rates of hydrogen production in biological systems based on the protein machinery of photosynthesis coupled with appropriate catalysts. We will also suggest areas for improvement and future directions for work in the field.

Engineering materials using supramolecular principles enables generalizable and modular platforms that have tunable chemical, mechanical, and biological properties. Applying this bottom‐up, molecular engineering‐based approach to therapeutic design affords unmatched control of emergent properties and functionalities. In preparing responsive materials for biomedical applications, the dynamic character of typical supramolecular interactions facilitates systems that can more rapidly sense and respond to specific stimuli through a fundamental change in material properties or characteristics, as compared to cases where covalent bonds must be overcome. Several supramolecular motifs have been evaluated toward the preparation of “smart” materials capable of sensing and responding to stimuli. Triggers of interest in designing materials for therapeutic use include applied external fields, environmental changes, biological actuators, applied mechanical loading, and modulation of relative binding affinities. In addition, multistimuli‐responsive routes can be realized that capture combinations of triggers for increased functionality. In sum, supramolecular engineering offers a highly functional strategy to prepare responsive materials. Future development and refinement of these approaches will improve precision in material formation and responsiveness, seek dynamic reciprocity in interactions with living biological systems, and improve spatiotemporal sensing of disease for better therapeutic deployment.

Azobenzene has been widely used as a photoregulator due to its reversible photoisomerization, large structural change between E and Z isomers, high photoisomerization yield, and high chemical stability. On the other hand, some azobenzene derivatives can be used as universal quenchers for many fluorophores. Nucleic acid is a good candidate to be modified because it is not only the template of gene expression but also widely used for building well-organized nanostructures and nanodevices. Because the size and polarity distribution of the azobenzene molecule is similar to a nucleobase pair, the introduction of azobenzene into nucleic acids has been shown to be an ingenious molecular design for constructing light-switching biosystems or light-driven nanomachines. Here we review recent advances in azobenzene-modified nucleic acids and their applications for artificial regulation of gene expression and enzymatic reactions, construction of photoresponsive nanostructures and nanodevices, molecular beacons, as well as obtaining structural information using the introduced azobenzene as an internal probe. In particular, nucleic acids bearing multiple azobenzenes can be used as a novel artificial nanomaterial with merits of high sequence specificity, regular duplex structure, and high photoregulation efficiency. The combination of functional groups with biomolecules may further advance the development of chemical biotechnology and biomolecular engineering.

Fungi are a prolific source of bioactive compounds, some of which have been developed as essential medicines and life-enhancing drugs. Genome sequencing has revealed that fungi have the potential to produce considerably more natural products (NPs) than are typically observed in the laboratory. Recently, there have been significant advances in the identification, understanding, and engineering of fungal biosynthetic gene clusters (BGCs). This review briefly describes examples of the engineering of fungal NP biosynthesis at the global, pathway, and enzyme level using in vivo and in vitro approaches and refers to the range and scale of heterologous expression systems available, developments in combinatorial biosynthesis, progress in understanding how fungal BGCs are regulated, and the applications of these novel biosynthetic enzymes as biocatalysts.

Si-Shen Feng Author for correspondence: Department of Chemical & Biomolecular Engineering, Department of Bioengineering, & Nanoscience & Nanotechnology Initiative (NUSNNI/NanoCore), National University of Singapore, Block E3, 05–29, 2 Engineering Drive 3, 117576, Singapore Tel.: +65 6576 3835 Fax: +65 6779 1936 chefess@nus.edu.sg Star-shaped block polymers as a molecular biomaterial for nanomedicine development

Synthetic Biology offers qualitatively new perspectives on the benefits of industrially harnessed biological processes. The ability to modify and reprogramme natural biology increases the scope of tailored bioprocesses and yields attractive prospects beyond conventional Biotechnology. The present review summarises the major achievements and categorises them according to a hierarchy of system levels. Similar structures are known in the engineering sciences and might prove useful for the future development of Synthetic Biology. The hierarchy encompasses several levels of detail. Biological (macro-)molecules present the most detailed level (parts), followed by compartmentalised or non-compartmentalised modules (devices). In the next level, parts and devices are combined into functional cells and further into cellular communities. The manifold interactions between biological entities of the same hierarchical level or between different levels are accounted for by networks, primarily metabolic pathways and regulatory circuits. Networks of different types are represented as a superordinate hierarchical level that achieves full system integration. On all these levels, extensive and sound scientific foundations exist regarding experimental but also theoretical methods. These have led to diverse manifestations of Synthetic Biology on the parts and devices levels. Investigations involving synthetic components on the systems scale represent the most difficult and remain limited in number. A main challenge lies with the quantitative prediction of interactions between different entities across different scales. Systems-theoretical approaches provide important tools to analyse complex biological behaviour and can support the design of artificial biological systems. A promising strategy is seen in an efficient modularisation that reduces biological systems to a limited set of functional modules with well-characterised interfaces. For the design of synthetic biological systems the interactions across these interfaces should be standardised to reduce complexity. Yet, the identification of modules and standardised interaction routes remains a non-trivial problem. Furthermore, an appropriate platform that efficiently describes replication and evolutionary processes has to be developed in order to extend the achievements of Synthetic Biology into designed biological processes.

A broad review of technologically focused work concerning biomolecules at interfaces is presented. The emphasis is on developments in interfacial biomolecular engineering that may have a practical impact in bioanalysis, tissue engineering, emulsion processing or bioseparations. We also review methods for fabrication in an attempt to draw out those approaches that may be useful for product manufacture, and briefly review methods for analysing the resulting interfacial nanostructures. From this review we conclude that the generation of knowledge and-innovation at the nanoscale far exceeds our ability to translate this innovation into practical outcomes addressing a market need, and that significant technological challenges exist. A particular challenge in this translation is to understand how the structural properties of biomolecules control the assembled architecture, which in turn defines product performance, and how this relationship is affected by the chosen manufacturing route. This structure-architecture-process-performance (SAPP) interaction problem is the familiar laboratory scale-up challenge in disguise. A further challenge will be to interpret biomolecular self- and directed-assembly reactions using tools of chemical reaction engineering, enabling rigorous manufacturing optimization of self-assembly laboratory techniques. We conclude that many of the technological problems facing this field are addressable using tools of modem chemical and biomolecular engineering, in conjunction with knowledge and skills from the underpinning sciences. (c) 2005 Elsevier Ltd. All rights reserved.

Biomolecular engineering is hot, the tech sector is turning around, and the United States will continue to dominate high-tech R&D in the coming decade. So say the IEEE Fellows in the second annual 2004 IEEE Technology Leaders Survey.

Protein drugs have revolutionized the pharmaceutical industry, offering new treatments for serious diseases. Since the first recombinant protein drug, Eli Lilly’s Humulin®, was approved 30 years ago (1), protein drugs have grown from esoteric specialty products to a major drug class. Of the 20 top selling drugs in the USA in the third quarter of 2012, nine are proteins. Despite these successes, the inherent instability of protein molecules remains an impediment to their development and to their safety and efficacy. One of the most serious types of instability is aggregation, the self-association of native protein through covalent and/or non-covalent interactions. Protein aggregates have been associated with increased or decreased drug potency and with an increased potential for immunogenic side effects, which can be life-threatening. In this themed issue of the AAPS Journal, we have assembled research and review articles that address the aggregation of therapeutic proteins. The issue was inspired by presentations at the 10th Annual Garnet E. Peck Symposium in Industrial Pharmacy, held at Purdue University in West Lafayette, Indiana, on October 11, 2012. At a practical level, interest in the aggregation of therapeutic proteins is driven by the need for stable formulations and robust manufacturing conditions. At the symposium, Dr. David Volkin (Dept. of Pharmaceutical Chemistry, University of Kansas) presented a series of case studies based on his recent work that address these issues. The cases showed the effects of excipients on the aggregation of an albumin fusion protein (2) and IgG2 monoclonal antibodies (mAbs) (3), the role of infusion bags in the solubility and aggregation of IgG4 mAbs (4), methods to ensure comparability during mAb process development (5), and an empirical phase diagram approach to identifying formulations that inhibit aggregation (6). In their review article in this special issue, Dr. Volkin and his coauthors summarize these case studies, present an overview of protein aggregation mechanisms, and describe high throughput approaches to monitoring protein stability (7). In developing drug products, the industry makes use of accelerated stability studies to estimate shelf life at the storage temperature based on degradation rates measured at higher temperatures. This extrapolation usually assumes that the temperature dependence of reaction rates follows Arrhenius behavior. However, protein aggregation often exhibits non-Arrhenius temperature dependence, even over relatively narrow temperature ranges. At the symposium, Dr. Chris Roberts (Dept. of Chemical and Biomolecular Engineering, University of Delaware) presented his group’s recent work on the mechanisms of protein aggregation and origins of non-Arrhenius behavior (8,9). In this themed issue, he and Dr. Wei Wang (Pfizer BioTherapeutics) summarize these mechanistic insights and discuss the implications for accelerated stability testing (10). Ensuring that protein drugs are aggregate-free requires robust, reproducible analytical methods. The ideal method would resolve aggregate and monomeric species, quantify aggregate size and concentration, and provide a low limit of detection, all while achieving high throughput at moderate cost. Current methods fall short of this ideal. For example, size exclusion chromatography (SEC) assays for soluble aggregates are expensive and low-throughput. Gel electrophoresis (e.g., SDS-PAGE) gives somewhat higher throughput but generally is not quantitative. To address these limitations, Dr. Mary Wirth (Dept. of Chemistry, Purdue University) and her colleagues are developing novel chromatographic materials based on silica colloidal crystals. These ordered arrays of silica particles provide plate heights in the low nanometer range, allowing for high resolution and rapid analysis times. For example, using a capillary packed with silica colloidal crystals and pressure-driven flow, the group has separated a monoclonal antibody and its aggregates in less than a minute with baseline resolution (11). Dr. Wirth and her coauthors summarize the group’s recent results in this themed issue (12). Together, these articles provide insight into our current understanding of protein aggregation. Gaps in fundamental understanding and applied technology remain, however, impeding our ability to monitor and inhibit aggregation in biologics. These gaps include limited understanding of the relative importance of partial unfolding, colloidal interactions, and chemical reaction in the protein aggregation process; a lack of understanding of the chemical and physical determinants of the immune response to protein drugs and their aggregated forms; poor agreement among available analytical methods for determining aggregate size and concentration, particularly in the subvisible range; and an incomplete understanding of the mechanistic effects of process stresses and formulation variables. We hope that the practical and theoretical perspectives assembled here will stimulate additional discussion and research in this important area.

The advances in high throughput screening technology for discovery of target molecules and the accumulation of functional genomics and proteomics data at an ever-accelerating rate will enable us to design and discover novel biomolecules and proteins on a rational basis in diverse areas of pharmaceutical, agricultural, industrial, and environmental applications. The biomolecular engineering will no doubt become one of the most important scientific disciplines in that it will enable us to comprehensively analyze gene expression patterns in both normal and diseased cells and to discover many new biologically active molecules rationally and systematically. As an applied molecular evolution technology, DNA shuffling will play a key role in biomolecular engineering. In contrast to the point mutation techniques, DNA shuffling exchanges large functional domains of sequences to search for the best candidate molecule, thus mimicking and accelerating the process of sexual recombination in the evolution of life. The phage-display system of combinatorial peptide libraries will be extensively exploited to design and create many more novel proteins, due to the relative ease of screening and identifying desirable proteins. Its application will be extended further into the science of protein–receptor or protein–ligand interactions. The bioinformatics including EST-based or SAGE-tag-based functional genomics and proteomics will continue to advance rapidly. Its biological knowledge base will expand the scope of biomolecular engineering, and the impact of well-coordinated biomolecular engineering research will be very significant on our understanding of gene expression, upregulation and downregulation, and posttranslational protein processing in healthy and diseased cells. The bioinformatics for genome and proteome analysis will contribute substantially toward ever more accelerated advances in pharmaceutical industry. When the functional genomics database, EST and SAGE techniques, microarray technique, and proteome analysis by 2-dimensional gel electrophoresis or capillary electrophoresis are all put to good use, the biomolecular engineering research will yield new drug discoveries, improved therapies, and new or significantly improved bioprocesses. With the advances in biomolecular engineering, the rate of finding new high-value peptides or proteins including antibodies, vaccines, enzymes, and therapeutic peptides will continue to be accelerated. The targets for rational design of biomolecules will be very broad, diverse, and complex, but many application goals can be achieved through the expansion of knowledge base on biomolecules of interest and their roles and functions in cells and tissues. In the near future, more therapeutic drugs and high-value biomolecules will be designed and produced for the treatment or prevention of not-so-easily-cured diseases such as cancers, genetic diseases, age-related diseases, and other metabolic diseases. Also anticipated are many more industrial enzymes that will be engineered to confer desirable properties for the process improvement and manufacturing of many high-value biomolecular products. Many more new metabolites including novel antibiotics that are active against resistant strains will be also produced by recombinant organisms having de novo engineered biosynthetic pathway enzyme systems. The biomolecular engineering era is here and a great deal of benefits can be derived from this field of scientific research for many years to come if we are willing to put it to good use.

Huimin Zhao is a professor of Chemical and Biomolecular Engineering at the University of Illinois at Urbana-Champaign and the holder of Centennial Chair. His research interests encompass many different aspects of synthetic biology and chemical biology. They include directed evolution and rational design of proteins and pathways, novel approaches for protein, pathway, and genome engineering, the molecular genetics and enzymology of natural product biosynthesis, the fundamental aspects of biocatalysis, and the use of protein engineering and metabolic engineering methods in designing platform organisms including E. coli and S. cerevisiae for the synthesis of important chemicals, antibiotics, and biofuels from renewable feedstocks.

Nanomedicine is an emerging field that integrates nanotechnology, biomolecular engineering, life sciences and medicine; it is expected to produce major breakthroughs in medical diagnostics and therapeutics. Due to the size-compatibility of nano-scale structures and devices with proteins and nucleic acids, the design, synthesis and application of nanoprobes, nanocarriers and nanomachines provide unprecedented opportunities for achieving a better control of biological processes, and drastic improvements in disease detection, therapy, and prevention. Recent advances in nanomedicine include the development of functional nanoparticle based molecular imaging probes, nano-structured materials as drug/gene carriers for in vivo delivery, and engineered molecular machines for treating single-gene disorders. This review focuses on the development of molecular imaging probes and engineered nucleases for nanomedicine, including quantum dot bioconjugates, quantum dot-fluorescent protein FRET probes, molecular beacons, magnetic and gold nanoparticle based imaging contrast agents, and the design and validation of zinc finger nucleases (ZFNs) and TAL effector nucleases (TALENs) for gene targeting. The challenges in translating nanomedicine approaches to clinical applications are discussed.

An interactive, multidisciplinary, public workshop, organized by a group of experts in biochemistry, biophysics, chemical and biomolecular engineering, chemistry, microbial metabolism, and protein structure and function, was held on January 6-7, 2011 in Washington, DC. Fundamental insights into the biological energy capture, storage, and transformation processes provided by speakers was featured in this workshop which included topics such as microbes living in extreme environments such as hydrothermal vents or caustic soda lakes (extremophiles) provided a fascinating basis for discussing the exploration and development of new energy systems. Breakout sessions and extended discussions among the multidisciplinary groups of participants in the workshop fostered information sharing and possible collaborations on future bioinspired research. Printed and web-based materials that summarize the committee's assessment of what transpired at the workshop were prepared to advance further understanding of fundamental chemical properties of biological systems within and between the disciplines. In addition, webbased materials (including two animated videos) were developed to make the workshop content more accessible to a broad audience of students and researchers working across disciplinary boundaries. Key workshop discussion topics included: Exploring and identifying novel organisms; Identifying patterns and conserved biological structures in nature; Exploring and identifying fundamental properties and mechanisms of knownmore » biological systems; Supporting current, and creating new, opportunities for interdisciplinary education, training, and outreach; and Applying knowledge from biology to create new devices and sustainable technology.« less

The biological production of fuels from renewable sources has been regarded as a feasible solution to the energy and environmental problems in the foreseeable future. Recently, the biofuel product spectrum has expanded from ethanol and fatty acid methyl esters (biodiesel) to other molecules, such as higher alcohols and alkanes, with more desirable fuel properties. In general, biosynthesis of these fuel molecules can be divided into two phases: carbon chain elongation and functional modification. In addition to natural fatty acid and isoprenoid chain elongation pathways, keto acid-based chain elongation followed by decarboxylation and reduction has been explored for higher alcohol production. Other issues such as metabolic balance, strain robustness, and industrial production process efficiency have also been addressed. These successes may provide both scientific insights into and practical applications toward the ultimate goal of sustainable fuel production.

Food materials science is an area of rapid growth in the larger field of food science. It is an applied area overlapping food science and food engineering that is concerned with structure, properties and processing of te materials used in food as well as their production and breakdown. The goals of researchers working in this area include: measuring the size and distribution of elements in a complex mixture, understanding how these elements interact and using this information to develop processes that will maximize the formation of useful structures. Examples are provided of newer methods for generating these structures such as the use of biomolecular engineering to design and produce novel proteins using amino acid sequences not found in nature. The concept of composite materials has been applied to foods and would appear to offer some advantages in dealing with these complex structures. An example is given dealing with the production of a food material composite, mixed and filled dairy gels, in which casein micelles are absorbed at the surface of protein-coated fat globules, leading to the formation of a copolymer network with overall greater strength. Of equal interest to the development of new food materials is how they behave under mechanical stress, and the idea of fracture behaviour, in particular notch sensitivity, is illustrated.

Department of Chemical & Biomolecular Engineering, National University of Singapore, 4 Engineering Drive 4, Singapore 117576, Singapore and Division of Bioengineering, National University of Singapore, 7 Engineering Drive 1, Singapore 117576, Singapore and Nanoscience & Nanotechnology Initiative, National University of Singapore, 2 Engineering Drive 3, Singapore 117587, Singapore Targeted nanomedicine for detection and treatment of circulating tumor cells

LEARNING OBJECTIVES After reading this chapter, you should: Understand the relationship between biomolecular engineering and chemical engineering. Understand the concepts of drug effectiveness and toxicity, the limitations of some of the simplest methods of drug administration, and the need for controlled delivery systems that optimize therapy for a particular drug. Understand how polymeric materials with different physical properties can be fashioned into matrix, microsphere, transdermal, and other delivery systems. Understand how tissue engineering has emerged as a possible solution for organ replacement or healing. Understand the biological significance of materials with nanoscale dimensions and how material scientists are learning to assemble materials that use or mimic biological principles of self-assembly. Prelude The early chapters of this book introduce some of the chemicals that are important in human biology. In fact, it is possible to think of the human body as an elaborate bag of chemicals. In the subspecialty called biomolecular engineering (or biotechnology), biomedical engineers examine the changes in chemical components within a biological system and develop methods for modifying these chemicals or their interactions. The concept of introducing chemicals to induce a change in a biological system is familiar; for example, we all have some experience with taking purified chemicals such as acetaminophen or ibuprofen as drugs to relieve pain. But new biological tools now make it possible to consider more complex chemical interventions such as gene therapy (in which a new deoxyribonucleic acid [DNA] sequence is introduced to allow expression of a new genetic activity).

Artificial chemistries are candidates for methodologies that model and design biochemical systems. If artificial chemistries can deal with such systems in beneficial ways, they may facilitate activities in the new area of biomolecular engineering. In order to explore such possibilities, we illustrate four models of biochemical pathways described in our artificial chemistry based on string pattern matching and recombination. The modeled pathways are the replication of DNA, transcription from DNA to mRNA, translation from mRNA to protein, and the oxidation of fatty acids. The descriptions show that the present approach has good modularity and scalability that will be useful for modeling a huge network of pathways. Moreover, we give a procedure to perform reasoning in the artificial chemistry, which checks whether a specified collection of molecules can be generated in a given model, and we demonstrate that it works on a model that describes a natural biochemical pathway.

This article gives short view on bioinformatics education at Keio university.

Due to the inability to form volatile etch products at temperatures less than 180 oC, the damascene process has been the prevailing patterning technology for copper. A simple, hydrogen (H2) plasma-based, low temperature etch process was developed to allow an alternative method to damascene technology. Cu thin films were etched by a H2 plasma in an inductively coupled plasma (ICP) reactor at temperatures below room temperature (10 oC). This process achieved anisotropic Cu features and an etch rate of ~13 nm/min. Ion bombardment was a contributor to Cu removal, since the etch rates were essentially proportional to the platen power with a constant coil power. However, lower etch rates of Cu were observed in an Ar plasma than in the H2 plasma, despite the fact that Ar is a more efficient sputter gas. Based on Cu etch rates and patterning results, the etch process involve both chemical and physical characteristics.

The last century witnessed spectacular advances in both microelectronics and biotechnology yet there was little synergy between the two. A challenge to their integration is that biological and electronic systems are constructed using divergent fabrication paradigms. Biology fabricates bottom-up with labile components, while microelectronic devices are fabricated top-down using methods that are 'bio-incompatible'. Biofabrication--the use of biological materials and mechanisms for construction--offers the opportunity to span these fabrication paradigms by providing convergent approaches for building the bio-device interface. Integral to biofabrication are stimuli-responsive materials (e.g. film-forming polysaccharides) that allow directed assembly under near physiological conditions in response to device-imposed signals. Biomolecular engineering, through recombinant technology, allows biological components to be endowed with information for assembly (e.g. encoded in a protein's amino acid sequence). Finally, self-assembly and enzymatic assembly provide the mechanisms for construction over a hierarchy of length scales. Here, we review recent advances in the use of biofabrication to build the bio-device interface. We anticipate that the biofabrication toolbox will expand over the next decade as more researchers enlist the unique construction capabilities of biology. Further, we look forward to observing the application of this toolbox to create devices that can better diagnose disease, detect pathogens and discover drugs. Finally, we expect that biofabrication will enable the effective interfacing of biology with electronics to create implantable devices for personalized and regenerative medicine.

On 9 and 10 October 1984 the European Communities' program in biomolecular engineering of crop plants and associated microorganisms met at the Carlsberg Laboratory in order to assess research progress, discuss future plans and coordinate efforts. Major trends and highlights apparent from the presentations are discussed here 1,2 .

The precise targeting of cells in deep tissues is one of the primary goals of nanomedicine. However, targeting a specific cellular population within an entire organism is challenging due to off-target effects and the need for deep tissue delivery. Focused ultrasound can reduce off-targeted effects by spatially restricting the delivery or action of molecular constructs to specific anatomical sites. Ultrasound can also increase the efficiency of nanotherapeutic delivery into deep tissues by enhancing the permeability of tissue boundaries, promoting convection, or depositing energy to actuate cellular activity. In this review we focus on the interface between biomolecular engineering and focused ultrasound, and describe the applications of this intersection in neuroscience, oncology, and synthetic biology. Ultrasound can be used to trigger the transport of therapeutic payloads into a range of tissues, including specific regions of the brain, where it can be targeted with millimeter precision through intact skull. Locally delivered molecular constructs can then control specific cells and molecular pathways within the targeted region. When combined with viral vectors and engineered neural receptors, this technique enables non-invasive control of specific circuits and behaviors. The penetrant energy of ultrasound can also be used to more directly actuate micro- and nanotherapeutic constructs, including microbubbles, vaporizable nanodroplets and polymeric nanocups, which nucleate cavitation upon ultrasound exposure, leading to local mechanical effects. In addition, it was recently discovered that a unique class of acoustic biomolecules – genetically encodable nanoscale protein structures called gas vesicles – can be acoustically “detonated” as sources of inertial cavitation. This enables the targeted disruption of selected cells within the area of insonation by gas vesicles that are engineered to bind cell surface receptors. It also facilitates ultrasound-triggered release of molecular payloads from engineered therapeutic cells heterologously expressing intracellular gas vesicles. Finally, focused ultrasound energy can be used to locally elevate tissue temperature and activate temperature-sensitive proteins and pathways. The elevation of temperature allows non-invasive control of gene expression in vivo in cells engineered to express thermal bioswitches. Overall, the intersection of biomolecular engineering, nanomaterials and focused ultrasound can provide unparalleled specificity in controlling, modulating and treating physiological processes in deep tissues.

This programme is essentially motivated by the need to allow the optimal exploitation by man of recent discoveries in modern biology and to stimulate in the Community v~e developments in applied fields where nations such as the U.S.A. and Japan have gained a considerable advance. Two main themes form ~he basis of the integrated research proposed for Community action. The first one deals with the development of the second generation of enzyme reactors, that is to say, with the exploitation of complex enzymatic reactions for the synthesis of elaborated products important to European industries. The second concerns the application of genetib engineering methods to organisms of importance for European agriculture and industry. Considerable attention is given here to the development of suitable host-vector systems and to the solution of the important practical problems which prevent the control of expression of foreign DNA.

The final contractors meeting of the Biomolecular Engineering programme in the sector “Second Generation Bioreactors” has taken place on 7–9 April in Compiegne. At this meeting the results were discussed and evaluated. The conclusions reached and the final reports are the basis for this review.

Cell factories have been largely exploited for the controlled production of substances of interest for food, pharma and biotech industries. Although human-controlled microbial production and transformation are much older, the cell factory concept was fully established in the 80’s through the intensive public and private investment. Strongly empowered by the then nascent recombinant DNA technologies and supported by the approval of recombinant insulin [1,2], the principle of controlled biological production as a convenient source of difficult-to-obtain molecules (especially those of high added value) deeply penetrated the industrial tissue, soon becoming a widespread platform aiming at cost-effective large-scale production [3]. The first generation cell factories (mainly composed by plain strains of the bacterium Escherichia coli and the yeast Saccharomyces cerevisae) were soon replaced by engineered variants. Resulting from the application of untargeted mutagenesis and phenotypic selection, conventional genetic modification, metabolic engineering, and more recently by systems metabolic engineering that integrates metabolic engineering with systems biology and synthetic biology, new strains having much enhanced performance have been progressively developed [4-19]. In parallel, mammalian and insect cells for the production of high quality proteins, and other microbial species appealing from an industrial point of view due to their unusual physiological traits, have been incorporated to the cell factory family [20,21]. This comprises algae, fungi, psychrophilic bacteria and moss, among others [22-26]. On the other hand, a set of food-grade lactic acid bacteria are under development as emerging platforms in food microbiology but also as a novel source of metabolites and proteins [27-34]. The physiological diversity of the microbial world offers an intricacy of biosynthetic pathways from which novel bio-products, including nano- or micro-structured materials [35-37], offer promises in even more diverse applications. Still, many substances and materials of industrial interest are nowadays produced by chemical synthesis, and the number of (recombinant) proteins approved for therapeutic use hardly reaches 200, a figure much lower than that initially presumed. However, both environmental concerns and medical and industrial needs strongly push towards a fully sustainable bio-production of a larger spectrum of substances. Then, how much limited is the economic feasibility of microbial production? Have the cell factories reached a plateau in their development? Is there more room for further exploitation of the microbial production? Systems metabolic engineering [38,39] offers a set of methodological and strategic tools for the design and optimization of metabolic and gene regulatory networks for the efficient production of chemicals (from pharmaceuticals to bulk chemicals and fuels) and materials (from plastics to high value materials) [5-7,40]. Furthermore, creation of new metabolic pathways and fine tuning of the existing ones have become possible [8,15]. This is already allowing the production of substances so far reluctant to microbial production under industrial requirements and under cost-effective processes, allowing us to overcome limitations of microbial cell factories. In silico genome-scale metabolic modeling and simulation are playing increasingly important roles in system-wide identification of target genes and pathways to be manipulated to enhance production of desired products. Profiling of transcriptome, proteome, metabolome, and/or fluxome is also becoming a routine practice in metabolic engineering for the better understanding of cellular characteristics under given genetic and/or environmental perturbations. Through the system-wide optimized design and development of microbial cell factories, many industrially important chemicals and materials could be efficiently produced. Some example products include, but not limited to, alcohols other than ethanol including propanol, butanol and isobutanol, dicarboxylic acids such as succinic acid, fumaric acid, malic acid and adipic acid, diols such as 1,3-propanediol, 1,2-propanediol, 1,4-butanediol and 2,3-butanediol, diamines such as putrescine and cadaverine, and even polymers including polyhydroxyalkanoates, polylactic acid, spider silk protein, poly-gamma-glutamic acid, and many others [40]. It is expected that increasingly diverse chemicals and materials that are currently produced by petrochemical industry will be produced by employing microbial cell factories. Improving cell factories by systems biotechnology to an industrially relevant level cannot be successfully reached by individual teams alone. A wide range of expertise is needed for the full development of a novel biotechnological process, including microbiology, chemistry, bioinformatics, biochemical engineering, but also agricultural and plant sciences and economics. After a proof of concept has shown the feasibility of a novel approach for cell factory engineering, large consortia or networks of fully skilled teams should take over in collaboration with Biotech companies to accomplish an economically feasible bioprocess. As an early example, the development of bacteria producing 1,3-propanediol from glucose has been well documented, and required the engineering of more than 70 genes, before process development and scale up were reasonably made [41]. Replacing materials which are produced from mineral oil by biobased products requires not only efficient technology but also the assessment of economical and ecological feasibility, as intensely studied for polylactic acid [42]. As the progress of industrial biotechnology did not accomplish the desired speed of development [43] academic researchers have teamed up with industry in publicly funded research centers worldwide. To name a few, the Joint BioEnergy Institute (JBEI) in the San Francisco Bay Area, the Novo Nordisk Foundation Center for Biosustainability (CFB) at the Technical University of Denmark, the Metabolic and Biomolecular Engineering Laboratory (MBEL) of the Korea Advanced Institute of Science and Technology (KAIST), the Dutch Klyuver Centre for Genomics of Industrial Fermentation, or the Austrian Centre of Industrial Biotechnology (ACIB) share the mission to speed up development of industrial bioprocesses by joining forces of different academic disciplines with industrial collaborators.