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Influence of culture aeration on the cellulase activity of Thermobifida fusca

  • Biotechnological Products and Process Engineering
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Abstract

Currently, one of the hurdles hindering efficient production of cellulosic biofuel is the recalcitrant nature of cellulose to hydrolysis. A wide variety of cellulase enzymes are found natively in microorganisms that can potentially be used to effectively hydrolyze cellulose to fermentable sugars. In this study, phenomenological and mechanistic parameters affecting cellulase activity were studied using the moderately thermophilic, aerobic, and cellulolytic microogainsm Thermobifida fusca. Two major sets of experiments were conducted to (1) study the mechanistic differences in growth in a flask compared to a bioreactor and (2) study the cell culture parameters influencing cellulase activity using a series of bioreactor experiments. Specific cellulase and specific endoglucanase activities were found to be higher in the bioreactor as compared to flask growth. Measurements of messenger RNA transcript levels of 18 cellulase-related genes and intracellular ATP levels indicated that measured enzyme activity was likely more influenced by post-transcriptional energetics rather than transcriptional regulation. By delineating the effects of culture aeration and stir speed using a bioreactor, it was found that cellulase activity increased with increasing aeration and increasing stir speeds (highest K l a) with a tradeoff of decreased cellular growth at the highest stir speeds tested (400 rpm). Overall, these results allude to a connection between aeration and oxidative respiration that lead to increased ATP allowing for increased cellulase synthesis as the primary constraint on overall cellulase activity.

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References

  • Ai YC, Wilson DB (2002) Mutation and expression of N233C–D506C of cellulase Cel6B from Thermobifida fusca in Escherichia coli. Enzyme Microb Technol 30:804–808

    Article  CAS  Google Scholar 

  • Bachmann SL, Mccarthy AJ (1991) Purification and cooperative activity of enzymes constituting the xylan-degrading system of Thermomonospora fusca. Appl Environ Microbiol 57:2121–2130

    CAS  Google Scholar 

  • Bandyopadhyay B, Humphrey AE, Taguchi H (1967) Dynamic measurement of the volumetric oxygen transfer coefficient in fermentation systems. Biotechnol Bioeng 9:533–544

    Article  CAS  Google Scholar 

  • Beki E, Nagy S, Vanderleyden J, Jager S, Kiss L, Fulop L, Hornok L, Kukolya J (2003) Cloning and heterologous expression of a beta-D-mannosidase (EC 3.2.1.25)-encoding gene from Thermobifida fusca TM51. Appl Environ Microbiol 69:1944–1952

    Article  CAS  Google Scholar 

  • Bevillard E, Goodheart DB, Karnati SKR, Bayer EA, Lamed R, Miron J, Nelson KE, Morrison M (2004) Ruminococcus albus 8 mutants defective in cellulose degradation are deficient in two processive endocellulases, Cel48A and Cel9B, both of which possess a novel modular architecture. J Bacteriol 186:136–145

    Article  Google Scholar 

  • Bradford MM (1976) Rapid and sensitive method for quantitation of microgram quantities of protein utilizing principal of protein-dye binding. Anal Biochem 72:248–254

    Article  CAS  Google Scholar 

  • Chen SL, Wilson DB (2007) Proteomic and transcriptomic analysis of extracellular proteins and mRNA levels in Thermobifida fusca grown on cellobiose and glucose. J Bacteriol 189:6260–6265

    Article  CAS  Google Scholar 

  • Demain AL, Newcomb M, Wu JHD (2005) Cellulase, clostridia, and ethanol. Microbiol Mol Biol Rev 69:124–154

    Article  CAS  Google Scholar 

  • Ferchak JD, Pye EK (1983) Effect of glucose and other sugars on the beta-1, 4-glucosidase activity of Thermomonospora fusca. Biotechnol Bioeng 25:2855–2864

    Article  CAS  Google Scholar 

  • Gusek TW, Johnson RD, Tyn MT, Kinsella JE (1991) Effect of agitational shear on growth and protease production by Thermomonospora fusca. Biotechnol Bioeng 37:371–374

    Article  CAS  Google Scholar 

  • Irwin DC, Zhang S, Wilson DB (2000) Cloning, expression and characterization of a Family 48 exocellulase, Cel48A, from Thermobifida fusca. Eur J Biochem 267:4988–4997

    Article  CAS  Google Scholar 

  • Jung H, Wilson DB, Walker LP (2003) Binding and reversibility of Thermobifida fusca Cel5A, Cel6B, and Cel48A and their respective catalytic domains to bacterial microcrystalline cellulose. Biotechnol Bioeng 84:151–159

    Article  CAS  Google Scholar 

  • Kim JH, Irwin D, Wilson DB (2004) Purification and characterization of Thermobifida fusca xylanase 10B. Can J Microbiol 50:835–843

    Article  CAS  Google Scholar 

  • Lao GF, Wilson DB (1996) Cloning, sequencing, and expression of a Thermomonospora fusca protease gene in Streptomyces lividans. Appl Environ Microbiol 62:4256–4259

    CAS  Google Scholar 

  • Lin E, Wilson DB (1988) Transcription of the CelE gene in Thermomonospora fusca. J Bacteriol 170:3838–3842

    CAS  Google Scholar 

  • Lykidis A, Mavromatis K, Ivanova N, Anderson I, Land M, DiBartolo G, Martinez M, Lapidus A, Lucas S, Copeland A, Richardson P, Wilson DB, Kyrpides N (2007) Genome sequence and analysis of the soil cellulolytic actinomycete Thermobifida fusca YX. J Bacteriol 189:2477–2486

    Article  CAS  Google Scholar 

  • OSullivan AC (1997) Cellulose: the structure slowly unravels. Cellulose 4:173–207

    Article  CAS  Google Scholar 

  • Saloheimo M, Lehtovaara P, Penttila M, Teeri TT, Stahlberg J, Johansson G, Pettersson G, Claeyssens M, Tomme P, Knowles JKC (1988) EGIII, a new endoglucanase from Trichoderma reesei: the characterization of both gene and enzyme. Gene 63:11–21

    Article  CAS  Google Scholar 

  • Spiridonov NA, Wilson DB (1998) Regulation of biosynthesis of individual cellulases in Thermomonospora fusca. J Bacteriol 180:3529–3532

    CAS  Google Scholar 

  • Spiridonov NA, Wilson DB (1999) Characterization and cloning of CelR, a transcriptional regulator of cellulase genes from Thermomonospora fusca. J Biol Chem 274:13127–13132

    Article  CAS  Google Scholar 

  • Spiridonov NA, Wilson DB (2001) Cloning and biochemical characterization of BglC, a beta-glucosidase from the cellulolytic actinomycete Thermobifida fusca. Curr Microbiol 42:295–301

    CAS  Google Scholar 

  • Watson DL, Wilson DB, Walker LP (2002) Synergism in binary mixtures of Thermobifida fusca cellulases Cel6B, Cel9A, and Cel5A on BMCC and Avicel. Appl Biochem Biotechnol 101:97–111

    Article  CAS  Google Scholar 

  • Wilson DB (2004) Studies of Thermobifida fusca plant cell wall degrading enzymes. Chem Rec 4:72–82

    Article  CAS  Google Scholar 

  • Xiao Z, Storms R, Tsang A (2004) Microplate-based filter paper assay to measure total cellulase activity. Biotechnol Bioeng 88:832–837

    Article  CAS  Google Scholar 

  • Zhang YHP, Himmel ME, Mielenz JR (2006) Outlook for cellulase improvement: screening and selection strategies. Biotechnol Adv 24:452–481

    Article  CAS  Google Scholar 

  • Zhou WL, Irwin DC, Escovar-Kousen J, Wilson DB (2004) Kinetic studies of Thermobifida fusca Cel9A active site mutant enzymes. Biochemistry 43:9655–9663

    Article  CAS  Google Scholar 

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  1. Department of Chemical and Life Science Engineering, Virginia Commonwealth University, 601 W. Main Street, Richmond, VA, 23284, USA

    Yu Deng & Stephen S. Fong

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  1. Yu Deng
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  2. Stephen S. Fong
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Correspondence to Stephen S. Fong.

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Deng, Y., Fong, S.S. Influence of culture aeration on the cellulase activity of Thermobifida fusca . Appl Microbiol Biotechnol 85, 965–974 (2010). https://doi.org/10.1007/s00253-009-2155-9

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  • DOI: https://doi.org/10.1007/s00253-009-2155-9

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