Biotechnology and Bioengineering

Cover image for Vol. 110 Issue 10

October 2013

Volume 110, Issue 10

Pages C1–C1, fmi–fmiv, vi–vi, 2563–2801

  1. Cover

    1. Top of page
    2. Cover
    3. Contents
    4. Spotlights
    5. Articles
    6. Communication to the Editors
    7. Communication to the Editor
    8. Communication to the Editors
  2. Contents

    1. Top of page
    2. Cover
    3. Contents
    4. Spotlights
    5. Articles
    6. Communication to the Editors
    7. Communication to the Editor
    8. Communication to the Editors
  3. Spotlights

    1. Top of page
    2. Cover
    3. Contents
    4. Spotlights
    5. Articles
    6. Communication to the Editors
    7. Communication to the Editor
    8. Communication to the Editors
  4. Articles

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

      Consensus engineering of sucrose phosphorylase: The outcome reflects the sequence input (pages 2563–2572)

      Dirk Aerts, Tom Verhaeghe, Henk-Jan Joosten, Gert Vriend, Wim Soetaert and Tom Desmet

      Version of Record online: 7 MAY 2013 | DOI: 10.1002/bit.24940

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      Consensus engineering is supposed to increase a protein's stability by recruiting residues that occur most frequently at the respective positions in a set of homologous sequences. However, the authors have found that the most abundant residues do not generate the most stable construct but one that displays an average stability. Whether this strategy is successful thus depends on which of the parent sequences is used as a point of comparison.

    2. In silico maturation of binding-specificity of DNA aptamers against Proteus mirabilis (pages 2573–2580)

      Nasa Savory, Danielle Lednor, Kaori Tsukakoshi, Koichi Abe, Wataru Yoshida, Stefano Ferri, Brian V Jones and Kazunori Ikebukuro

      Version of Record online: 22 APR 2013 | DOI: 10.1002/bit.24922

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      The authors developed DNA aptamers against Proteus mirabilis by the successful approach combining whole-cell based systematic evolution of ligands by exponential enrichment (Cell-SELEX) and specificity improvement by in silico maturation (ISM). P. mirabilis is a prominent cause of catheter-associated urinary tract infections (CAUTIs) among patients undergoing long-term bladder catheterization. Aptamers that specifically recognize P. mirabilis would be highly valuable not only in biosensor development for rapid diagnosis, but also as a tool for studying membrane-associated proteins in this organism.

    3. Surface engineering of a cutinase from Thermobifida cellulosilytica for improved polyester hydrolysis (pages 2581–2590)

      Enrique Herrero Acero, Doris Ribitsch, Anita Dellacher, Sabine Zitzenbacher, Annemarie Marold, Georg Steinkellner, Karl Gruber, Helmut Schwab and Georg M. Guebitz

      Version of Record online: 29 APR 2013 | DOI: 10.1002/bit.24930

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      Cutinases from Thermobifida sp. have been recognized as powerful tools for hydrolysis of polyethylene terephthalate (PET). In order to investigate the role of amino acids located on the surface of cutinases on PET hydrolysis, selected amino acids of the low active cutinase 2 from T. cellulosilytica were exchanged by amino acids of the highly active cutinase 1 using site-directed mutagenesis. The results clearly demonstrated that surface properties like amino acids located outside the active site on the protein surface play an important role in PET hydrolysis.

    4. Production of a novel O-methyl-isoflavone by regioselective sequential hydroxylation and O-methylation reactions in Streptomyces avermitilis host system (pages 2591–2599)

      Kwon-Young Choi, EunOk Jung, Yung-Hun Yang and Byung-Gee Kim

      Version of Record online: 26 APR 2013 | DOI: 10.1002/bit.24931

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      The authors have developed a recombinant S. avermitilis strain producing a novel O-methyl-isoflavone using selected OMTs and SAM synthetase genes. The 3′-OMD produced by the SeOMT3 strain is a novel O-methyl-isoflavone which is not a naturally occurring compound. Also this is the first report that overexpression of metK induces the production of secondary metabolites through oxidation reactions followed by a subsequent O-methylation reaction.

    5. Biofuels and Environmental Biotechnology

      Shearing of biofilms enables selective layer based microbial sampling and analysis (pages 2600–2605)

      Yang Lu, Frances Slater, Ricardo Bello-Mendoza and Damien J. Batstone

      Version of Record online: 16 MAY 2013 | DOI: 10.1002/bit.24947

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      A method to remove microbes selectively from successive spatial layers through hydraulic shearing is demonstrated on anaerobic granules. This could also be used in conjunction with community profiling methods to combine the depth of microbial community profiling with localization capability. Dominant population shifted from Bacteroidetes and Anaerolinea in outer layers to Syntrophomonas and Geobacter in inner layers. It may also provide opportunity to conduct further live culture analysis by extracting whole live organisms.

    6. Consolidated bioprocessing of highly concentrated jerusalem artichoke tubers for simultaneous saccharification and ethanol fermentation (pages 2606–2615)

      Lihao Guo, Jian Zhang, Fengxian Hu, Dewey Dy Ryu and Jie Bao

      Version of Record online: 22 APR 2013 | DOI: 10.1002/bit.24929

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      A practical CBP process technology in the helical ribbon bioreactor for economically ethanol production using highly concentrated Jat is provided according to the scheme shown in the Figure. The high level of ethanol yield contributes to Saccharomyces cerevisiae DQ1, which produces relatively large amounts of inulinase, and the helical ribbon stirring bioreactor, which provides good mixing performance under high Jat loading.

    7. Evolutionary engineering of Saccharomyces cerevisiae for enhanced tolerance to hydrolysates of lignocellulosic biomass (pages 2616–2623)

      María P. Almario, Luis H. Reyes and Katy C. Kao

      Version of Record online: 11 JUL 2013 | DOI: 10.1002/bit.24938

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      The evolutionary dynamics of Saccharomyces cerevisiae during in vitro evolution in hydrolysates of lignocellulosic biomass are described. The adaptive evolution method visualizing evolution in real time (VERT) was used for the directed evolution of S. cerevisiae for enhanced tolerance to hydrolysates. VERT was used to facilitate the isolation of adaptive mutants from evolving population and ramp-up of selective pressure.

    8. Bioprocess Engineering and Supporting Technologies

      Industrial symbiosis: Corn ethanol fermentation, hydrothermal carbonization, and anaerobic digestion (pages 2624–2632)

      Brandon M. Wood, Lindsey R. Jader, Frederick J. Schendel, Nicholas J. Hahn, Kenneth J. Valentas, Patrick J. McNamara, Paige M. Novak and Steven M. Heilmann

      Version of Record online: 26 APR 2013 | DOI: 10.1002/bit.24924

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      Conventional dry-grind corn ethanol production is controversial due to energy and water usage as well as economic issues. To address these concerns hydrothermal carbonization coupled with anaerobic digestion of stillage intermediates created in corn ethanol manufacturing have been examined as alternative downstream processing operations. This novel combination substantially reduced downstream energy consumption, treated process water, and generated fatty acid and hydrochar products.

    9. Evaluating differences of metabolic performances: Statistical methods and their application to animal cell cultivations (pages 2633–2642)

      O. Hädicke, V. Lohr, Y. Genzel, U. Reichl and S. Klamt

      Version of Record online: 30 APR 2013 | DOI: 10.1002/bit.24926

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      Monitoring and analyzing metabolic key parameters, such as metabolic rates, is routinely applied in cell culture process development to demonstrate specific advantages of one experimental setup over another. However, a systematic assessment of whether the observed differences are statistically significant is often missing. This work provides a guideline for statistical analyses with well-established methods to avoid false-positive results in comparative cultivation studies. Using a case study, implications of different sources of variations on the significance of observed differences are analyzed systematically.

    10. Optimized extract preparation methods and reaction conditions for improved yeast cell-free protein synthesis (pages 2643–2654)

      C. Eric Hodgman and Michael C. Jewett

      Version of Record online: 7 JUL 2013 | DOI: 10.1002/bit.24942

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      Cell-free protein synthesis is a powerful technology for simple and efficient protein production. Here, the authors describe a novel CFPS platform derived from Saccharomyces cerevisiae extract. By developing a streamlined crude extract preparation protocol and optimizing the CFPS reaction conditions Hodgman and Jewett achieved active firefly luciferase synthesis yields of 7.7 ± 0.5 μg mL−1. Their results set the stage for developing a yeast CFPS platform for high-yielding and cost-effective expression of a variety of protein therapeutics and protein libraries.

    11. Bioseparations and Downstream Processing

      A tandem laboratory scale protein purification process using Protein A affinity and anion exchange chromatography operated in a weak partitioning mode (pages 2655–2663)

      Michael Shamashkin, Ranga Godavarti, Timothy Iskra and Jon Coffman

      Version of Record online: 29 MAY 2013 | DOI: 10.1002/bit.24955

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      Manufacturing of high titer mAbs in older facilities can generate large volume in-process pools exceeding the capacity of older storage vessels. This paper describes a feasibility study of a bench-scale tandem process for the purification of mAbs employing three in-line steps (Protein A chromatography, flow-through anion-exchange chromatography, and virus filtration) without the need to collect in-process pools. Numerous challenges related to in-line neutralization of Protein A elution were explored.

    12. High-throughput screening for the development of a monoclonal antibody affinity precipitation step using ELP-z stimuli responsive biopolymers (pages 2664–2676)

      Rahul D. Sheth, Bhawna Madan, Wilfred Chen and Steven M. Cramer

      Version of Record online: 16 MAY 2013 | DOI: 10.1002/bit.24945

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      A high-throughput screening strategy was employed to develop a robust affinity precipitation process for Monoclonal antibody monoclonal antibody purification using, Elastin-like polypeptide fused to binding domain, ELP-Z. mAb yield and aggregation were monitored as a function of the operating conditions. The results suggest that a room temperature process providing high mAb yields with low aggregation is possible, demonstrating its suitability as a potential mAb capture process.

    13. Cellular and Metabolic Engineering

      Using the cre–lox system to randomize target gene expression states and generate diverse phenotypes (pages 2677–2686)

      Bradley Niesner and Narendra Maheshri

      Version of Record online: 3 JUN 2013 | DOI: 10.1002/bit.24952

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      By flanking promoters in budding yeast with inverted loxP sites and adding Cre recombinase to randomize their orientation, the authors demonstrate the ability to produce a combinatorial explosion (2N, 3N or 4N) of expression states with a small (N) number of genetic modifications. They show how this approach can reveal novel epistatic interactions in base excision repair and suggest how it may be deployed for biotechnological applications.

    14. Enhanced degradation of haloacid by heterologous expression in related Burkholderia species (pages 2687–2696)

      Xianbin Su, Liyu Deng, Ka Fai Kong and Jimmy S. H. Tsang

      Version of Record online: 22 APR 2013 | DOI: 10.1002/bit.24917

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      Bacterium Burkholderia species MBA4 produced a dehalogenase that transforms haloacid to utilizable substrate. Around a hundred units of enzyme were produced in the presence of haloacid. The gene encoding for the dehalogenase was cloned and transformed to B. caribensis, B. phymatum and B. xenovorans and enable these cells to grow on haloacid. Two to three hundred units of dehalogenase were produced in these cells without haloacid and up to 670 units of enzyme were found in mono-chloroacetate-grown cells.

    15. Investigating contactless high frequency ultrasound microbeam stimulation for determination of invasion potential of breast cancer cells (pages 2697–2705)

      Jae Youn Hwang, Nan Sook Lee, Changyang Lee, Kwok Ho Lam, Hyung Ham Kim, Jonghye Woo, Ming-Yi Lin, Kassandra Kisler, Hojong Choi, Qifa Zhou, Robert H. Chow and K. Kirk Shung

      Version of Record online: 22 APR 2013 | DOI: 10.1002/bit.24923

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      The authors investigate the application ofcontactless high frequency ultrasound microbeam stimulation(HFUMS) for determining the invasion potential of breast cancer cells. Chow and coworkers found that HFUMS elicits significant calcium elevation in highly invasive breast cancer cells compared to weakly invasive breast cancer cells. Thus, this finding suggest that HFUMS may serve as a novel tool to determine the invasion potential of breast cancer cells, and with further refinement may offer a rapid test for invasiveness of tumor biopsies in situ.

    16. A human embryonic stem cell line adapted for high throughput screening (pages 2706–2716)

      Nicolas J. Caron, Blair K. Gage, Michael D. O'Connor, Connie J. Eaves, Timothy J. Kieffer and James M. Piret

      Version of Record online: 29 APR 2013 | DOI: 10.1002/bit.24936

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      CA1S hESCs exhibit improved plating efficiency and homogeneity over classic hESC lines. This attribute leads to highly uniform seeding, growth, and differentiation without loss of pluripotency. Thus CA1S cells are suitable for high throughput screening studies focused on optimizing culture, differentiation and development of hESCs.

    17. Enhanced FK506 production in Streptomyces tsukubaensis by rational feeding strategies based on comparative metabolic profiling analysis (pages 2717–2730)

      Menglei Xia, Di Huang, Shanshan Li, Jianping Wen, Xiaoqiang Jia and Yunlin Chen

      Version of Record online: 16 MAY 2013 | DOI: 10.1002/bit.24941

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      The present work provided a valuable method for debottlenecking low-yield microbial natural product. Based on comparative metabolic profiling analysis, the metabolites associated closely with FK506 production were identified. By in-depth analysis on the metabolic relevance between these key metabolites and FK506 biosynthesis, vital regulation points for FK506 overproduction were uncovered. Rational feeding strategies were then proposed which finally resulted in a 61.4% increase in target product.

    18. Engineering Science of Biological Systems

      Cell spreading and proliferation in response to the composition and mechanics of engineered fibrillar extracellular matrices (pages 2731–2741)

      Antony K. Chen, Frank W. Delrio, Alexander W. Peterson, Koo-Hyun Chung, Kiran Bhadiraju and Anne L. Plant

      Version of Record online: 22 APR 2013 | DOI: 10.1002/bit.24921

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      Chen and coworkers aimed to study the effect of different extracellular matrix (ECM) compositions and mechanical properties on cell response. They engineered novel thin films of ECM proteins consisting of fibrillar Type-1 collagen (COL) and fibronectin (FN) that allowed systematic examination of the effects of matrix composition and mechanics on cell spreading and proliferation. This work should aid in understanding the relative contributions of multiple ECM factors on cell behaviors in the complex in vivo environment.

    19. Electrostatic properties of confluent Caco-2 cell layer correlates to their microvilli growth and determines underlying transcellular flow (pages 2742–2748)

      P. Vandrangi, D.D. Lo, R. Kozaka, N. Ozaki, N. Carvajal and V.G.J. Rodgers

      Version of Record online: 27 JUN 2013 | DOI: 10.1002/bit.24939

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      The authors analyzed the cellular morphology, transport characteristics, and the claudin 4 expression for Caco-2 cells; The decrease in the measured normal zeta potential was attributed to the observed microvilli developed on their apical surface.

    20. Systems Biotechnology

      Long-term adaptation of Saccharomyces cerevisiae to the burden of recombinant insulin production (pages 2749–2763)

      Ali Kazemi Seresht, Ana Luisa Cruz, Erik de Hulster, Marit Hebly, Eva Akke Palmqvist, Walter van Gulik, Jean-Marc Daran, Jack Pronk and Lisbeth Olsson

      Version of Record online: 14 MAY 2013 | DOI: 10.1002/bit.24927

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      A data-driven approach, consisting of metabolome and transcriptome analysis, was applied to study the secretory production of human insulin in S. cerevisiae in prolonged chemostat cultivations. As part of the long-term adaptation of the cells towards the burden of insulin production, a metabolic re-modelling of the cells was observed, leading to a decrease insulin expression capacity of the cells. The presented information about the crosstalk between the different metabolic pathways represents a basic foundation for targeted strain engineering strategies.

    21. The challenge of improved secretory production of active pharmaceutical ingredients in Saccharomyces cerevisiae: A case study on human insulin analogs (pages 2764–2774)

      Ali Kazemi Seresht, Eva A. Palmqvist, Gerd Schluckebier, Ingrid Pettersson and Lisbeth Olsson

      Version of Record online: 26 APR 2013 | DOI: 10.1002/bit.24928

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      Secretion of two human insulin analog precursors (IAPs) with minor differences in their amino acid sequences and a more than seven fold difference in secretion yield was investigated. Global transcriptome analysis carried out in chemostat experiments identified distinct steps during the protein maturation pathway to be differentially regulated, and indicated an increased degradation of the IAP with the low secretion yield. In silico protein structure modelling of the IAPs suggested a difference in conformational stability, induced by the amino acid substitution, which most likely contributed to the difference in trafficking through the secretory pathway and thus the large difference in secretion yields.

    22. Tissue Engineering and Delivery Systems

      Electrospun aligned PHBV/collagen nanofibers as substrates for nerve tissue engineering (pages 2775–2784)

      Molamma P. Prabhakaran, Elham Vatankhah and Seeram Ramakrishna

      Version of Record online: 29 APR 2013 | DOI: 10.1002/bit.24937

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      Electrospun nanofibers of PHBV/Collagen have huge potential as scaffolds for nerve tissue engineering, due to their biocompatibility and slow degradation properties. Composite PHBV/Collagen nanofibers improved cell growth and differentiation of PC12 cells, and the current studies demonstrated the ability of cells to extend their neurite on aligned PHBV/Collagen nanofibers along the direction of orientation of the fibers, providing contact guidance for enhanced cellular alignment and suggesting the nano-topographical significance of scaffolds suitable for nerve tissue regeneration.

  5. Communication to the Editors

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

      Protein cell-surface display through in situ enzymatic modification of proteins with a poly(Ethylene glycol)-lipid (pages 2785–2789)

      Urara Tomita, Satoshi Yamaguchi, Yasukazu Maeda, Kazuki Chujo, Kosuke Minamihata and Teruyuki Nagamune

      Version of Record online: 26 APR 2013 | DOI: 10.1002/bit.24933

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      The authors we report a facile enzymatic method for displaying target proteins on living cells via in situ enzymatic site-specific modification of a poly(ethylene glycol)(PEG)-lipid. Using this method, the desired amount of active target proteins can be reliably incorporated onto cell membranes within a few hours. This method is a promising tool for regulating and reinforcing cell-cell interactions in cell and tissue engineering fields.

  6. Communication to the Editor

    1. Top of page
    2. Cover
    3. Contents
    4. Spotlights
    5. Articles
    6. Communication to the Editors
    7. Communication to the Editor
    8. Communication to the Editors
    1. Cellular and Metabolic Engineering

      Butyrate production in engineered Escherichia coli with synthetic scaffolds (pages 2790–2794)

      Jang-Mi Baek, Suman Mazumdar, Sang-Woo Lee, Moo-Young Jung, Jae-Hyung Lim, Sang-Woo Seo, Gyoo-Yeol Jung and Min-Kyu Oh

      Version of Record online: 22 APR 2013 | DOI: 10.1002/bit.24925

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      When the three exogenous enzymes of the butyrate pathway were spatially organized by scaffold proteins, the metabolically engineered E. coli efficiently re-directed carbon flux to butyrate and dramatically increased its productivity.

  7. Communication to the Editors

    1. Top of page
    2. Cover
    3. Contents
    4. Spotlights
    5. Articles
    6. Communication to the Editors
    7. Communication to the Editor
    8. Communication to the Editors
    1. Tissue Engineering and Delivery Systems

      Translocation of cell penetrating peptides on Chlamydomonas reinhardtii (pages 2795–2801)

      Arumuganainar Suresh and Yeu-Chun Kim

      Version of Record online: 30 APR 2013 | DOI: 10.1002/bit.24935

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      Genetic engineering of microalgal cells, while considered a promising field of biotechnological research, is inhibited mainly by the difficulty in delivering macromolecules through the cell wall and plasma membrane. In this study, Suresh and Kim identified a cell penetrating peptide (CPP) that can efficiently translocate into microalgae, namely pVEC. The results of this study indicate that pVEC is an attractive candidate that can be employed as a peptide delivery vector in microalgae engineering. It is expected that the establishment of pVEC as a genetic engineering tool will open an array of new directions for biological and industrial applications.

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