Biotechnology and Bioengineering

Cover image for Vol. 110 Issue 7

July 2013

Volume 110, Issue 7

Pages C1–C1, fmi–fmvi, 1811–2062

  1. Cover

    1. Top of page
    2. Cover
    3. Contents
    4. Spotlights
    5. Reviews
    6. Articles
    7. Communications to the Editor
    1. You have free access to this content
      Biotechnology and Bioengineering: Volume 110, Number 7, July 2013 (page C1)

      Article first published online: 28 MAY 2013 | DOI: 10.1002/bit.24674

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      Cover Legend Escherichia coli was metabolically engineered to produce fumaric acid, an important industrial platform chemical, by reinforcing the oxidative TCA cycle fl ux under aerobic condition. Image created by Sang Yup Lee and Chan Woo Song at KAIST, Korea.

  2. Contents

    1. Top of page
    2. Cover
    3. Contents
    4. Spotlights
    5. Reviews
    6. Articles
    7. Communications to the Editor
  3. Spotlights

    1. Top of page
    2. Cover
    3. Contents
    4. Spotlights
    5. Reviews
    6. Articles
    7. Communications to the Editor
  4. Reviews

    1. Top of page
    2. Cover
    3. Contents
    4. Spotlights
    5. Reviews
    6. Articles
    7. Communications to the Editor
    1. Transcription activator-like effector nucleases (TALENs): A highly efficient and versatile tool for genome editing (pages 1811–1821)

      Ning Sun and Huimin Zhao

      Article first published online: 7 APR 2013 | DOI: 10.1002/bit.24890

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      Transcription activator-like effector nucleases (TALENs) have recently emerged as a revolutionary genome editing tool in many different organisms and cell types. In this article, the authors comprehensively review the development of TALEN technology in terms of scaffold optimization, DNA recognition, and repeat array assembly. In addition, they provide some perspectives on the future development of this technology.

  5. Articles

    1. Top of page
    2. Cover
    3. Contents
    4. Spotlights
    5. Reviews
    6. Articles
    7. Communications to the Editor
    1. ArticlesBiocatalysis, Protein Engineering, and Nanobiotechnology

      Effects of short elastin-like peptides on filamentous particles and their transition behavior (pages 1822–1830)

      Adam P. Hathorne and Harry Bermudez

      Article first published online: 22 FEB 2013 | DOI: 10.1002/bit.24854

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      Using a protein engineering approach, the authors have created filamentous phage particles decorated with short elastin-like polypeptides (ELPs). Characterization of these particles reveals changes in surface packing and secondary structure, although the particles cannot be thermally triggered to macroscopically aggregate. Evidence from analogous ELP peptides suggests that their microscopic switching behavior is preserved even at very short lengths. These results highlight important considerations for ELP functionality in nanoscale environments.

    2. Biocatalysis, Protein Engineering, and Nanobiotechnology

      Influence of outer membrane c-type cytochromes on particle size and activity of extracellular nanoparticles produced by Shewanella oneidensis (pages 1831–1837)

      Chun Kiat Ng, Krishnakumar Sivakumar, Xin Liu, Munusamy Madhaiyan, Lianghui Ji, Liang Yang, Chuyang Tang, Hao Song, Staffan Kjelleberg and Bin Cao

      Article first published online: 1 MAR 2013 | DOI: 10.1002/bit.24856

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      Extracellular biogenic silver and silver sulfide nanoparticles produced by the Shewanella oneidensis mutant lacking outer membrane cytochromes MtrC and OmcA are smaller and show higher bactericidal and catalytic activity, respectively, compared to the wild type. Our results suggest that it may be possible to control particle size and activity of extracellular biogenic nanoparticles via controlled expression of the genes encoding surface proteins. In addition, the need for novel methods that ensure efficient retention of extracellular nanoparticles in biosynthesis processes is also underlined.

    3. Biochemical characterization of an alcohol dehydrogenase from Ralstonia sp. (pages 1838–1848)

      Justyna Kulig, Amina Frese, Wolfgang Kroutil, Martina Pohl and Dörte Rother

      Article first published online: 22 FEB 2013 | DOI: 10.1002/bit.24857

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      A unique alcohol dehydrogenase from Ralstonia sp. (RADH) accepting bulky substrates was biochemically characterized. The enzyme's stability was improved by employment of Ca2+ ions. RADH revealed satisfactory stability and excellent biocatalytical properties at mild conditions (pH and temperature).

    4. Engineering of a bacterial tyrosinase for improved catalytic efficiency towards D-tyrosine using random and site directed mutagenesis approaches (pages 1849–1857)

      Susan Molloy, Jasmina Nikodinovic-Runic, Leona B. Martin, Hermann Hartmann, Francisco Solano, Heinz Decker and Kevin E. O'Connor

      Article first published online: 22 FEB 2013 | DOI: 10.1002/bit.24859

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      Bacterial tyrosinases generally exhibit poor activity towards D-tyrosine in comparison to L-tyrosine. Through in vitro mutagenesis Ralstonia solanacearum tyrosinase variants with between 2.8- and 16-fold higher catalytic efficiency for D-tyrosine were obtained. Amino acid changes occurred outside the active site with T183I and N322S improving affinity while V153A improved catalytic activity but decreased affinity of the enzyme for D-tyrosine. Engineered tyrosinases contain amino acid changes affecting D-tyrosine more than L-tyrosine oxidation.

    5. Introduction of selective intersubunit disulfide bonds into self-assembly protein scaffold to enhance an artificial multienzyme complex's activity (pages 1858–1864)

      Hidehiko Hirakawa, Ayano Kakitani and Teruyuki Nagamune

      Article first published online: 22 FEB 2013 | DOI: 10.1002/bit.24861

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      Sulfolobus solfataricus proliferating cell nuclear antigen (PCNA) is a ring-shaped symmetric heterotrimer. Multiple enzymes can be co-localized on the PCNA ring by fusing them to the C-termini of the three PCNA subunits. Selective intersubunit disulfide bond formation by a natural oxidizing reagent successfully stabilized the multienzyme complex to fulfill its potential.

    6. Periplasmic expression of carbonic anhydrase in Escherichia coli: A new biocatalyst for CO2 hydration (pages 1865–1873)

      Tushar N. Patel, Ah-Hyung Alissa Park and Scott Banta

      Article first published online: 1 MAR 2013 | DOI: 10.1002/bit.24863

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      Two isoforms of carbonic anhydrase (CA) were periplasmically expressed to generate whole-cell biocatalysts for CO2 hydration. The catalysts exhibited increased thermal stability, a favorable consequence of protein immobilization. The cost of the catalyst may be reduced since protein purification is not required. However, their activities were decreased relative to their corresponding purified enzymes, an expected result from the introduction of a transport barrier. Finally, the catalysts can mediate carbon mineralization, a permanent carbon capture and sequestration technology in which CO2 is converted to a carbonate salt.

    7. Biofuels and Environmental Biotechnology

      You have free access to this content
      Engineered thermostable fungal Cel6A and Cel7A cellobiohydrolases hydrolyze cellulose efficiently at elevated temperatures (pages 1874–1883)

      Indira Wu and Frances H. Arnold

      Article first published online: 1 MAR 2013 | DOI: 10.1002/bit.24864

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      High enzyme costs remain a major hurdle for the conversion of cellulosic feedstocks into fermentable sugars for microbial biofuel production. Wu and Arnold showed that enhancing the thermostability of a fungal Cel6A cellobiohydrolase is an effective strategy to reduce cost by increasing hydrolysis yield or by reducing hydrolysis time. A binary mixture of engineered thermostable Cel6A and Cel7A retains wild-type like synergy and achieves higher hydrolysis yield than the equivalent wild-type mixture at the respective optimum temperatures.

    8. A modified metabolic model for mixed culture fermentation with energy conserving electron bifurcation reaction and metabolite transport energy (pages 1884–1894)

      Fang Zhang, Yan Zhang, Man Chen, Mark C.M. van Loosdrecht and Raymond J. Zeng

      Article first published online: 22 FEB 2013 | DOI: 10.1002/bit.24855

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      A modified metabolic model of mixed culture fermentation (MCF) with two new assumptions: energy conserving electron bifurcation reaction and metabolite transport energy, is proposed to improve the Rodriguez model. The modeling results in a continuous stirred tank reactor show that the metabolite distribution is consistent with the literature, which demonstrates our modified model is more realistic than previous proposed model concepts. It also indicates that inclusion of these two new assumptions for MCF is sound from the viewpoint of biochemistry and biophysiology.

    9. Renewable bio ionic liquids-water mixtures-mediated selective removal of lignin from rice straw: Visualization of changes in composition and cell wall structure (pages 1895–1902)

      Xue-Dan Hou, Ning Li and Min-Hua Zong

      Article first published online: 22 FEB 2013 | DOI: 10.1002/bit.24862

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      [Ch][AA] ILs-water mixtures proved to be highly promising solvents for lignocellulosic biomass pretreatment under mild conditions. Good sugar yields were obtained in the enzymatic hydrolysis of rice straw after pretreatment using these mixtures at 90°C. The changes in the contents and distribution of the main components in cell walls were visualized by microscopic techniques.

    10. A novel nonwoven hybrid bioreactor (NWHBR) for enhancing simultaneous nitrification and denitrification (pages 1903–1912)

      Fangang Meng, Yuan Wang, Li-Nan Huang, Jie Li, Feng Jiang, Shiyu Li and Guang-Hao Chen

      Article first published online: 4 MAR 2013 | DOI: 10.1002/bit.24866

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      In this study, a novel nonwoven hybrid bioreactor (NWHBR) was developed for SND process, in which the nonwoven part played dual roles as biofilm carrier and created membrane-like separation of flocculent sludge. The authors results showed that the mass transfer within the biofilm and thus the nitrogen removal rates in the reactor were enhanced as a result of permeate drag. In addition, the NWHBR had low physical back-washing frequency and low air-to-water ratio.

    11. Bioprocess Engineering and Supporting Technologies

      Induced effect of Na+ on ganoderic acid biosynthesis in static liquid culture of Ganoderma lucidum via calcineurin signal transduction (pages 1913–1923)

      Yi-Ning Xu, Xiao-Xia Xia and Jian-Jiang Zhong

      Article first published online: 22 FEB 2013 | DOI: 10.1002/bit.24852

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      Ganoderma lucidum, a traditional Chinese medicinal mushroom, produces an anti-tumor triterpenoid—ganoderic acid (GA). Na+ addition to the static liquid fermentation increased the transcriptional levels of genes of Ca2+ sensors and GA biosynthesis by triggering calcineurin signals, and greatly enhanced the GA production. The work provides a simple and efficient induction strategy to improve GA production and demonstrates the regulatory mechanism of Na+ on the GA biosynthesis through calcineurin signaling transduction.

    12. Identifying analytics for high throughput bioprocess development studies (pages 1924–1935)

      Spyridon Konstantinidis, Simyee Kong and Nigel Titchener-Hooker

      Article first published online: 15 FEB 2013 | DOI: 10.1002/bit.24850

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      High throughput (HT) studies are best facilitated by analytical methods suited to deployment in this mode. The HT-capability is governed by features of each method. These features, however, need to be considered collectively, not on a stand-alone basis, and within the context of the planned experimental study so as to identify those analytics which are fit for purpose. The authors describe such an approach and discuss its application for a wide selection of analytical methods, frequently employed during bioprocess development activities.

    13. Remote delivery of hydroxyl radicals via secondary chemistry of a nonthermal plasma effluent (pages 1936–1944)

      S. Reed Plimpton, Mark Gołkowski, Deborah G. Mitchell, Chad Austin, Sandra S. Eaton, Gareth R. Eaton, Czeslaw Gołkowski and Martin Voskuil

      Article first published online: 22 FEB 2013 | DOI: 10.1002/bit.24853

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      A non-thermal plasma device enables remote delivery of short-lived hydroxyl radicals in an air stream.

    14. Software sensor design considering oscillating conditions as present in industrial scale fed-batch cultivations (pages 1945–1955)

      V. Lyubenova, S. Junne, M. Ignatova and P. Neubauer

      Article first published online: 26 MAR 2013 | DOI: 10.1002/bit.24870

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      A method is described for monitoring of industrial-scale fed-batch processes with a step-wise software sensor approach composed of a cascade structure. Process kinetics are simulated considering time-varying yield coefficients. The software sensor design is realized after a linear transformation using a logarithmic function. The effectiveness of the proposed monitoring approach is demonstrated with experimental data of biomass, oxygen consumption and the respiratory quotient of Bacillus subtilis fed-batch cultivations in a two-compartment scale-down reactor.

    15. Bioseparations and Downstream Processing

      Protein crystallization in stirred systems—scale-up via the maximum local energy dissipation (pages 1956–1963)

      Benjamin Smejkal, Bernhard Helk, Jean-Michel Rondeau, Sabine Anton, Angelika Wilke, Peter Scheyerer, Jacqueline Fries, Dariusch Hekmat and Dirk Weuster-Botz

      Article first published online: 9 FEB 2013 | DOI: 10.1002/bit.24845

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      Technical-scale protein crystallization in stirred vessels can be a viable alternative to purification as well as polishing via preparative chromatography, particularly for therapeutic proteins. However, little systematic know-how exists. In this study, scale-up of protein crystallization was investigated using a Fab-fragment of the therapeutic antibody Canakinumab and lysozyme. A faster onset of crystallization was observed at an optimum stirrer speed and a comparable crystallization behavior was achieved by applying a constant maximum local energy dissipation for scale-up.

    16. Clarification of recombinant proteins from high cell density mammalian cell culture systems using new improved depth filters (pages 1964–1972)

      Nripen Singh, Kara Pizzelli, Jonathan K. Romero, James Chrostowski, Greg Evangelist, James Hamzik, Neil Soice and K.S. Cheng

      Article first published online: 7 MAR 2013 | DOI: 10.1002/bit.24848

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      This article describes novel filter designs which result in improved primary and secondary clarification for flocculated high cell density mammalian cell culture systems. It discusses the use of new Clarisolve™ depth filters with higher dirt holding capacity to achieve a centrifuge-free clarification train and process compression with high product recovery at a manufacturing scale of 2000 L. Implementing the strategies discussed will improve yield, lower operating costs and reduce the process footprint.

    17. An ultra scale-down characterization of low shear stress primary recovery stages to enhance selectivity of fusion protein recovery from its molecular variants (pages 1973–1983)

      Eduardo C. Lau, Simyee Kong, Shaun McNulty, Claire Entwisle, Ann Mcilgorm, Kate A. Dalton and Mike Hoare

      Article first published online: 16 MAR 2013 | DOI: 10.1002/bit.24865

      The bioprocessing of an Fc-fusion protein from mammalian cell culture broths is studied with particular focus on selective recovery in early clarification stages. The challenge is to understand the process engineering issues underlying the extent of selectivity achieved especially with respect to undesired variants of the protein and to non-protein contaminants. The use of ultra scale-down methods is used to explore process options and identify the importance of controlling the of the cell culture broth to levels of high shear stress if good selectivity is to be achieved.

    18. Using high throughput screening to define virus clearance by chromatography resins (pages 1984–1994)

      Lisa Connell-Crowley, Elizabeth A. Larimore and Ron Gillespie

      Article first published online: 26 MAR 2013 | DOI: 10.1002/bit.24869

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      High throughput screening (HTS) in a 96-well batch binding format was used to map xMuLV removal on Capto adhere and Q Sepharose Fast Flow. Results indicate that NaCl concentration and impurity level, but not pH, are key parameters that can impact xMuLV binding. HTS results compared favorably to data generated in the column format. Integrating HTS-derived virus binding information into downstream process development allows for chromatography development that maximizes selectivity of contaminants while ensuring acceptable virus reduction capabilities.

    19. Cellular and Metabolic Engineering

      Highly efficient production of the Alzheimer's γ-Secretase integral membrane protease complex by a multi-gene stable integration approach (pages 1995–2005)

      Jean-René Alattia, Mattia Matasci, Mitko Dimitrov, Lorène Aeschbach, Sowmya Balasubramanian, David L. Hacker, Florian M. Wurm and Patrick C. Fraering

      Article first published online: 15 FEB 2013 | DOI: 10.1002/bit.24851

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      A transposon-based multi-gene integration system was used to generate recombinant Chinese hamster ovary cell lines co-expressing stably the four subunits (NCT, Aph1, Pen2 and PS1) of the Alzheimer's γ-secretase integral membrane protease complex. Ten-liter suspension cultures of selected clones allowed the production of milligram amounts of highly purified and active γ-secretase. The authors conclude that transposon-based multigene transfer is a powerful approach for the production of highly purified multiprotein complexes, including γ-secretase, for structure determination and drug discovery applications.

    20. Knockout of pgdS and ggt genes improves γ-PGA yield in B. subtilis (pages 2006–2012)

      Viola Scoffone, Daniele Dondi, Ginevra Biino, Giovanni Borghese, Dario Pasini, Alessandro Galizzi and Cinzia Calvio

      Article first published online: 15 FEB 2013 | DOI: 10.1002/bit.24846

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      γ-PGA is a very promising biopolymer produced by Bacillus species, with a multiplicity of potential applications. However, for industrial use, bacterial productivity must be increased. In a B. subtilis producer strain, the removal of two enzymes (PgdS and GGT), involved in polymer degradation, significantly improves polymer recovery and doubles productivity without affecting bacterial growth.

    21. Peak antibody production is associated with increased oxidative metabolism in an industrially relevant fed-batch CHO cell culture (pages 2013–2024)

      Neil Templeton, Jason Dean, Pranhitha Reddy and Jamey D. Young

      Article first published online: 4 MAR 2013 | DOI: 10.1002/bit.24858

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      The authors performed 13C metabolic flux analysis (MFA) upon an industrially relevant CHO fed-batch culture and analyzed four separate phases of growth. Peak growth was characterized by a highly glycolytic metabolic state, corresponding with minimal antibody production and maximal lactate production. In contrast, the authors determined that a highly oxidative state of metabolism corresponded with peak specific antibody production.

    22. Metabolic engineering of Escherichia coli for the production of fumaric acid (pages 2025–2034)

      Chan Woo Song, Dong In Kim, Sol Choi, Jae Won Jang and Sang Yup Lee

      Article first published online: 1 MAR 2013 | DOI: 10.1002/bit.24868

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      Through rational stepwise metabolic engineering strategies coupled with in silico flux response simulation, the authors developed an engineered E. coli strain capable of producing 28.2 g/L fumaric acid with the overall yield and productivity of 0.389 g fumaric acid/g glucose and 0.448 g/L/h, respectively, in 63 h by aerobic fed-batch culture.

    23. Tissue Engineering and Delivery Systems

      Improved recellularization of ex vivo vascular scaffolds using directed transport gradients to modulate ECM remodeling (pages 2035–2045)

      Zehra Tosun and Peter S. McFetridge

      Article first published online: 29 APR 2013 | DOI: 10.1002/bit.24934

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      The aim of the present work was to create a directed growth environment that allows cells to fully populate an ex vivo-derived vascular scaffold and maintain viability over extended periods. The influence of directed nutrient gradients where precisely controlled perfusion conditions significantly affects cell migration, distribution and function, resulting in pronounced effects on construct mechanics during early remodeling events, is described.

    24. Dynamic compressive loading differentially regulates chondrocyte anabolic and catabolic activity with age (pages 2046–2057)

      Nikki. L. Farnsworth, Lorena R. Antunez and Stephanie J. Bryant

      Article first published online: 1 MAR 2013 | DOI: 10.1002/bit.24860

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      Dynamic loading has emerged as an important regulator of chondrocytes and a critical component for tissue engineering cartilage. This research demonstrates that the age of the donor from which chondrocytes are isolated differentially regulates how chondrocytes respond to loading, altering both anabolic and catabolic responses. Overall, this study suggests that the design of a cartilage tissue engineering strategy may need to be tailored to the age of the donor.

  6. Communications to the Editor

    1. Top of page
    2. Cover
    3. Contents
    4. Spotlights
    5. Reviews
    6. Articles
    7. Communications to the Editor
    1. Biocatalysis, Protein Engineering, and Nanobiotechnology

      Reducing infectivity of HIV upon exposure to surfaces coated with N,N-dodecyl, methyl-polyethylenimine (pages 2058–2062)

      Stephen E. Gerrard, Alyssa M. Larson, Alexander M. Klibanov, Nigel K.H. Slater, Carl V. Hanson, Barbara F. Abrams and Mary K. Morris

      Article first published online: 7 APR 2013 | DOI: 10.1002/bit.24867

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      The infectivity of high-titer, cell-free HIV in culture media and human milk is rapidly reduced upon exposure to polyethylene slides painted with the linear hydrophobic polycation N, N-dodecyl,methyl-polyethylenimine (DMPEI). Accompanying viral p24 protein and free viral RNA analysis of solutions exposed to DMPEI-coated surfaces suggest that virion attachment to the polycationic surface and its subsequent inactivation are the likely mechanism of this phenomenon.

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