Abstract
In this study, d-amino acid oxidase (DAAO) and catalase (CAT) in the permeabilized recombinant Pichia pastori cells were well investigated. It appeared that their thermal stability was negatively correlated with the apparent enzymatic activities. The frozen-melted cells presented the best stability and the lowest apparent activities of DAAO and CAT, whereas the cetyltrimethylammonium bromide (CTAB) permeabilized cells displayed the weakest stability and the highest apparent activities of the two enzymes. Simultaneous action of DAAO and CAT in the CTAB-permeabilized cells and glutaryl-7-aminocephalosporanic acid acylase (GA) immobilized on carrier contributed to the conversion of cephalosporin C (CPC) to 7-aminocephalosporanic acid (7-ACA) with a yield of 76.2%. During such a reaction cycle, no visible activity loss occurred at the immobilized GA, whereas the loss rates of DAAO and CAT activities were about 0.029 and 1.13 U min−1, respectively. Nevertheless, this problem could be easily solved by continuous feeding of the new permeabilized cell suspension at the rate of 6 ml h−1 to the reactor. Following such a fed-batch strategy, these permeabilized cells and the immobilized GA could be efficiently reused for 6 and 15 reaction cycles, respectively, yielding around 76% 7-ACA at each reaction cycle.
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References
Aail P, Bále V, Michalková E (1995) Biotransformation of cephalosporin C to 7-aminocephalosporanic acid with coimmobilized biocatalyst in a batch bioreactor. Bioproc Biosyst Eng 12:249–252
Aail P, Bále V, Michalková E (1996) Biotransformation of cephalosporin C to 7-aminocephalosporanic acid with coimmobilized biocatalyst in a batch bioreact. Bioproc Biosyst Eng 14:105–109
Aebi H (1984) Assay of catalase in vitro conditions. In: Parker L (ed) Methods in enzymology 105. New York, Academic Press, pp 122–127
Bachhawat Nandita, Gowda LR, Bhat SG (1996) Single step method of preparation of detergent-permeabilized kluyveromyces fragilis for lactose hydrolysis. Process Biochem 31:21–25
Becka S, Skrob F, Plhackova K, Kujan P, Holler P, Kyslik P (2003) Cross-linked cell aggregates of Trigonopsis variabilis: d-amino acid oxidase catalyst for oxidation of cephalosporin C. Biotechnol Let 25:227–233
Betancor L, Hidalgo A, Fernandez-Lorente R, Mateo C, Rodriguez V, Fuentes M, Lopez-Gallego F, Fernandez-Lafuente R, Guisan JM (2003) Use of physicochemical tools to determine the choice of optimal enzyme: stabilization of d-amino oxidase. Biotechnol Prog 19:784–788
Bianchi D, Bortolo R, Golini P, Cesti P (1998) Enzymatic transformation of cephalosporin C to 7-ACA by simultaneous action of immobilized d-amino acid oxidase and glutaryl-7-ACA acylase. Appl Biochem Biotechnol 73:257–268
Brodelius P, Nisson K, Mosbach K (1981) Production of α-keto acids: immobilized cells of Trigonopsis variabilis containing d-amino acid oxidase. Appl Biochem Biotechnol 6:293–308
Gough S, Deshpande M, Scher M, Rosazza JPN (2001) Permeabilization of Pichia pastoris for glycolate oxidase activity. Biotechnol Lett 23:1535–1537
Gowda LR, Bachhawat N, Bhat SG (1991) Permeabilization of Bakers’ yeast by cetyitrimethylammonium bromide for intracellular enzyme catalysis. Enzyme Microb Technol 13:154–157
Lopez-Gallego F, Batencor L, Hidalgo A, Mateo C, Fernandez-Lafuente R, Guisan JM (2005) One-pot conversion of cephalosporin C to 7-aminocephalosporanic acid in the absence of hydrogen peroxide. Adv Synth Catal 347:1804–1810. doi:10.1002/adsc.200505099
Lopez-Gallego F, Batencor L, Sio CF, Reis CR, Jimenez PN, Guisan JM, Quax WJ, Fernandez-Lafuente R (2008) Evaluation of different glutaryl acylase mutants to improve the hydolysis of cephalosporin C in the absence of hydrogen peroxide. Adv Synth Catal 350:343–348. doi:10.1002/adsc.200700320
Luo H, Yu HM, Li Q, Shen ZY (2004) Cloning and co-expression of d-amino acid oxidase and glutaryl-7-ACA acylase genes in Escherichia coli. Enzyme Microb Technol 35:514–518. doi:10.1016/j.enzmictec.2004.08.036
Matsuda A, atsuyama K, Yamamoto K, Ichikawa S, Komatsu KI (1987) Cloning and characterization of the genes for two distinct cephalosporin acylases from a Pseudomonas strain. J Bacteriol 58:5815–5820
Monti D, Carrea G, Riva S, Baldaro E, Frara G (2002) Characterication of an industrial biocatalyst: immobilized glutaryl-7-ACA acyalse. Biotechnol Bioeng 70:239–244
Moreno JA, Ruiz CA, Catalan J, Galan MA, Chem J (2004) Thermal deactivation and inhibition of d-amino acid oxidase in permeabilized cells of the yeast Trigonopsis variabilis. J Chem Technol Biotechnol 79:321–326. doi:10.1002/jctb.954
Nigam VK, Kundu S, Ghosh P (2005) Single-step conversion of cephalosporin C to 7-aminocephalosporanic acid by free and immobilized cells of Pseudomonas diminuta. Appl Biochem Biotechnol 126:13–21
Oh B, Kim K, Yoon J, Chung K, Shin Y, Lee D, Kim Y (2003) Deacylation activity of cephalosporin acylase to cephalosporin C is improved by changing the side-chain conformations of active-site residues. Biochem Biophys Res Commun 310:19–27. doi:10.1016/j.bbrc.2003.08.110
Oh B, Kim K, Park J, Yoon J, Han D, Kim Y (2004) Modifying the substrate specificity of penicillin G acylase to cephalosporin acylase by mutating active-site residues. Biochem Biophys Res Commun 319:486–492. doi:10.1016/j.bbrc.2004.05.017
Parmar A, Kumar H, Marwaha SS, Kennedy JF (1998) Recent trends in enzymatic conversion of cephalosporin C to 7-aminocephalosporanic acid (7-ACA). Crit Rev Biotechnol 18:1–12
Pilone M, Pollegioni L (2002) d-amino acid oxidase as an industrial biocatalyst. Biocatal Biotransform 20:145–159. doi:10.1080/10242420290020679
Pollegioni L, Lorenzi S, Rosini E, Marcone GL, Molla G, Verga R, Cabri W, Pilone MS (2005) Evolution of an acylase active on cephalosporin C. Protein Sci 14:3064–3076. doi:10.1110/ps.051671705
Shibuya Y, Matsumoto K, Fujii T (1981) Isolation and properties of 7-β-(4-carboxybutanamide)-cephalosporinic acid acylase-producing bacteria. Agric Biol Chem 45:1561–1567
Tan Q, Song QS, Wei DZ (2006) Single-pot conversion of cephalosporin C to 7-aminocephalosporanic acid using cell-bound and support-bound enzymes. Enzyme Microb Technol 39:1166–1172. doi:10.1016/j.enzmictec.2006.02.028
Tan Q, Song QS, Zhang YW, Wei DZ (2007) Characterization and application of d-amino acid oxidase and catalase within permeabilized Pichia pastoris cells in bioconversions. Appl Biochem Biotechnol 136:279–290
Upadhya R, Nagajyothi H, Bhat SD (1999) d-Amino acid oxides and catalase of detergent permeabilized Rhodotorula gracilis cells and its potential use for the synthesis of α-keto acids. Process Biochem 35:7–13
Upadhya R, Nagajyothi H, Bhat SD (2000) Stabilization of d-amino acid oxidase and catalase in permeabilized Rhodotorula gracilis cells and its application for the preparation of α-keto acids. Biotechnol Bioeng 68:430–436
Vicenzi JT, Hansen GJ (1993) Enzymatic oxidation of cephalosporin C using whole cells of the yeast Trigonopsis variabilis within a “cross-flow filter-reactor”. Enzyme Microb Technol 15:281–285
Wolf KF, Koller KP, Lange G, Liesum A, Sauber K, Schreuder H, Aretz W, Kabsch W (2002) Structure-based prediction of modification in glutarylamidase to allow single-step enzymatic production of 7-aminocephalosporanic acid from cephalosporin C. Protein Sci 11:92–103. doi:10.1101/ps.27502
Yu J, Li DY, Zhang YJ, Yang S, Li RB, Yuan ZY (2002) High expression of Trigonopsis variabilis d-amino acid oxidase in Pichia pastoris. J Mol Catal B Enzym 18:291–297
Zheng HB, Zhu TB, Chen J, Zhao YH, Jiang WH, Zhao GP, Yang S, Yang YL (2007) Construction of recombinant Escherichia coli D11/pMSTO and its use in enzymatic preparation of 7-aminocephalosporanic acid in one pot. J Biotechnol 129:400–405. doi:10.1016/j.jbiotec.2007.01.021
Zhu TB, Shen J, Zhang YF, Yang YL, Jiao RS (2001) Permeabilization of yeast Trigonopsis variabilis FA10 cells for enhancing apparent activity of d-amino acid oxidase. Chin J Biotechnol 17:73–77
Acknowledgments
We thank Professor Zhongyi Yuan for kindly providing the recombinant P. pastoris strains. We are also grateful to Dr Tianwen Wang for reading the manuscript. This study was supported by the Key Disciplinary Foundation of Shanghai.
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Tan, Q., Zhang, Y., Song, Q. et al. Single-pot conversion of cephalosporin C to 7-aminocephalosporanic acid in the absence of hydrogen peroxide. World J Microbiol Biotechnol 26, 145–152 (2010). https://doi.org/10.1007/s11274-009-0153-9
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DOI: https://doi.org/10.1007/s11274-009-0153-9