- Email: [email protected]
Reactivation of biocatalysts partially inactivated by organic solvent
S92
Abstracts / Journal of Biotechnology 131S (2007) S74–S97
Table 1 Kinetic parameters
diminishing the detrimental effects of the cosubstrate, in the biotransformation stage a fed-batch cultivation system was used.
Substrate
Vmax (g/L)
Km (g/L)
Kig (g/L)
Kss (g/L)
Ksn (g/L)
GF GF2 GF3 GF3
89.9 31.4 18.7 8.5
197.9 186.8 256.5 308.1
78.0 40.2 44.7 –
785.0 – – –
– – – 784.6
doi:10.1016/j.jbiotec.2007.07.158 31. Reactivation of biocatalysts partially inactivated by organic solvent Lorena Wilson ∗ , Oscar Romero, Jos´e Vergara, Andr´es Illanes School of Biochemical Engineering, Pontificia Universidad Cat´olica de Valpara´ıso, Valpara´ıso, Chile E-mail address: [email protected] (L. Wilson).
Fig. 1. Model prediction and experimental data (symbols).
References Duan, K.J., Chen, J.S., Sheu, D.C., 1994. Enzyme Microb. Technol. 16, 334– 339. Hernalsteens, S., 2006. Doutorado. Universidade Estadual de Campinas, S˜ao Paulo. Jung, K.H., Yun, J.W., Kang, K.R., Lim, J.Y., 1989. Enzyme Mircob. Technol. 11, 491–494. Remaud-Simeon, M., Lopez-Minguia, A., Pelenc, V., Paul, F., Monsan, P., 1994. Appl. Biochem. Biotechnol. 44 (2), 101–117.
Biocatalyst reactivation may have a profound impact in production systems in highly denaturant conditions. This is the case in the synthesis of some -lactam antibiotics, where the use of the cosolvent is necessary to displace the equilibrium towards synthesis, or selectively depress the competing hydrolytic reactions. Two penicillin acylases (PGA) were used: cross-linked enzyme aggregates (PGA-CLEA) and multi-point immobilized PGA in glyoxyl agarose (PGA-GA). These biocatalysts were incubated at 14 ◦ C at two dioxane concentrations (30 and 70%, v/v). Biocatalysts were inactivated down to different residual activities and then re-incubated in aqueous medium to promote reactivation, studying the effect of temperature (5, 14 and 25 ◦ C) and ionic strength (10% (w/v) ammonium sulphate). Best conditions were obtained at 25 ◦ C in the absence of salt. PGA-GA was more stable and exhibited higher levels of reactivation than PGA-CLEA, reaching 113% of the initial activity in the water-dioxane medium. Stability of PGA-GA was also studied in repeated batches of cephalosporin G synthesis to evaluate the option of reactivation and reutilization with intermediate stages of reactivation. The strategy proved useful: high levels of reactivation (from 45 to 85%) were obtained and conversion yield was maintained over 80%.
doi:10.1016/j.jbiotec.2007.07.157 30. Optically active alcohols formation by yeast biotransformation Ovidiu Popa a,∗ , Narcisa Babeanu b , Adrian Vamanu a , Manuel Drugulescu c , Emanuel Vamanu b a
Research Centre for Applied Biochemistry and Biotechnology, Centrul Cercetari Biochimie Aplicata si Biotehnologie BIOTEHNOL, 59 Marasti Bld., 011464 Bucharest, Romania b Faculty of Biotechnology, University of Agronomical Sciences and Veterinary Medicine, Bucharest, Romania c Compreserv, Bucharest, Romania E-mail address: ovid [email protected]. Several strains of the genera Saccharomyces, Candida and Hansenula were investigated for their ability to produce lphenyl-acetyl-carbinol. As the lethal effect of benzaldehyde on the living cells is well known, the selected strain of Hansenula polymorpha was adapted to 3.5 g/L benzaldehyde. The pH and cell age effects on the level of L-PAC was investigated. For
Acknowledgement Work funded by CONICYT Grant N◦ 1070361. doi:10.1016/j.jbiotec.2007.07.159 32. Enantioselective enzymatic carboligation using thiamine diphosphate dependent enzymes in a solid/gas bioreactor Antje Spiess a,∗ , Renaud Mikolajek a , Martina Pohl b , Sylvain Lamare c a
RWTH Aachen University, 52074 Aachen, Germany J¨ulich, J¨ulich, Germany c Universit´ e de La Rochelle, La Rochelle, Germany b FZ
E-mail address: [email protected]. Enzymatic gas phase catalysis, where gaseous substrates are converted by dry enzymes to gaseous products, has been proven to increase operational stability of enzymes at high space-time yields (Trivedi et al., 2006). The carboligation using thiamine
Abstracts / Journal of Biotechnology 131S (2007) S74–S97
Table 1 Kinetic parameters
diminishing the detrimental effects of the cosubstrate, in the biotransformation stage a fed-batch cultivation system was used.
Substrate
Vmax (g/L)
Km (g/L)
Kig (g/L)
Kss (g/L)
Ksn (g/L)
GF GF2 GF3 GF3
89.9 31.4 18.7 8.5
197.9 186.8 256.5 308.1
78.0 40.2 44.7 –
785.0 – – –
– – – 784.6
doi:10.1016/j.jbiotec.2007.07.158 31. Reactivation of biocatalysts partially inactivated by organic solvent Lorena Wilson ∗ , Oscar Romero, Jos´e Vergara, Andr´es Illanes School of Biochemical Engineering, Pontificia Universidad Cat´olica de Valpara´ıso, Valpara´ıso, Chile E-mail address: [email protected] (L. Wilson).
Fig. 1. Model prediction and experimental data (symbols).
References Duan, K.J., Chen, J.S., Sheu, D.C., 1994. Enzyme Microb. Technol. 16, 334– 339. Hernalsteens, S., 2006. Doutorado. Universidade Estadual de Campinas, S˜ao Paulo. Jung, K.H., Yun, J.W., Kang, K.R., Lim, J.Y., 1989. Enzyme Mircob. Technol. 11, 491–494. Remaud-Simeon, M., Lopez-Minguia, A., Pelenc, V., Paul, F., Monsan, P., 1994. Appl. Biochem. Biotechnol. 44 (2), 101–117.
Biocatalyst reactivation may have a profound impact in production systems in highly denaturant conditions. This is the case in the synthesis of some -lactam antibiotics, where the use of the cosolvent is necessary to displace the equilibrium towards synthesis, or selectively depress the competing hydrolytic reactions. Two penicillin acylases (PGA) were used: cross-linked enzyme aggregates (PGA-CLEA) and multi-point immobilized PGA in glyoxyl agarose (PGA-GA). These biocatalysts were incubated at 14 ◦ C at two dioxane concentrations (30 and 70%, v/v). Biocatalysts were inactivated down to different residual activities and then re-incubated in aqueous medium to promote reactivation, studying the effect of temperature (5, 14 and 25 ◦ C) and ionic strength (10% (w/v) ammonium sulphate). Best conditions were obtained at 25 ◦ C in the absence of salt. PGA-GA was more stable and exhibited higher levels of reactivation than PGA-CLEA, reaching 113% of the initial activity in the water-dioxane medium. Stability of PGA-GA was also studied in repeated batches of cephalosporin G synthesis to evaluate the option of reactivation and reutilization with intermediate stages of reactivation. The strategy proved useful: high levels of reactivation (from 45 to 85%) were obtained and conversion yield was maintained over 80%.
doi:10.1016/j.jbiotec.2007.07.157 30. Optically active alcohols formation by yeast biotransformation Ovidiu Popa a,∗ , Narcisa Babeanu b , Adrian Vamanu a , Manuel Drugulescu c , Emanuel Vamanu b a
Research Centre for Applied Biochemistry and Biotechnology, Centrul Cercetari Biochimie Aplicata si Biotehnologie BIOTEHNOL, 59 Marasti Bld., 011464 Bucharest, Romania b Faculty of Biotechnology, University of Agronomical Sciences and Veterinary Medicine, Bucharest, Romania c Compreserv, Bucharest, Romania E-mail address: ovid [email protected]. Several strains of the genera Saccharomyces, Candida and Hansenula were investigated for their ability to produce lphenyl-acetyl-carbinol. As the lethal effect of benzaldehyde on the living cells is well known, the selected strain of Hansenula polymorpha was adapted to 3.5 g/L benzaldehyde. The pH and cell age effects on the level of L-PAC was investigated. For
Acknowledgement Work funded by CONICYT Grant N◦ 1070361. doi:10.1016/j.jbiotec.2007.07.159 32. Enantioselective enzymatic carboligation using thiamine diphosphate dependent enzymes in a solid/gas bioreactor Antje Spiess a,∗ , Renaud Mikolajek a , Martina Pohl b , Sylvain Lamare c a
RWTH Aachen University, 52074 Aachen, Germany J¨ulich, J¨ulich, Germany c Universit´ e de La Rochelle, La Rochelle, Germany b FZ
E-mail address: [email protected]. Enzymatic gas phase catalysis, where gaseous substrates are converted by dry enzymes to gaseous products, has been proven to increase operational stability of enzymes at high space-time yields (Trivedi et al., 2006). The carboligation using thiamine