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Purification and Characterization of a Low Molecular Weight of β-Mannanase from Penicillium occitanis Pol6

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Abstract

The highest β-mannanase activity was produced by Penicillium occitanis Pol6 on flour of carob seed, whereas starch-containing medium gave lower enzymes titles. The low molecular weight enzyme was purified to homogeneity by ammonium sulfate precipitation, gel filtration, and ion-exchange chromatography procedures. The purified β-mannanase (ManIII) has been identified as a glycoprotein (carbohydrate content 5%) with an apparent molecular mass of 18 kDa. It was active at 40 °C and pH 4.0. It was stable for 30 min at 70 °C and has a broad pH stability (2.0–12.0). ManIII showed K m, V max, and K cat values of 17.94 mg/ml, 93.52 U/mg, and 28.13 s−1 with locust bean gum as substrate, respectively. It was inhibited by mannose with a K I of 0.610−3 mg/ml. ManIII was activated by CuSO4 and CaCl2 (2.5 mM). However, in presence of 2.5 mM Co2+, its activity dropped to 60% of the initial activity. Both N-terminal and internal amino acid sequences of ManIII presented no homology with mannanases of glycosides hydrolases. During incubation with locust bean gum and Ivory nut mannan, the enzyme released mainly mannotetraose, mannotriose, and mannobiose.

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Acknowledgments

This research was financially supported in part by “Ministére de la recheche scientifique, de la Technologie et du dévelopement de compétences”. We express our gratitude to Prof. G. Tiraby and Dr. H. Durand (Cayla Company, France) for kindly supplying the P. occitanis (Pol 6) strain used in this work. We are also grateful to Prof. Radhouane Ellouz, Prof. Hamadi Attia, Prof. Youssef Gargouri, Prof. Mongi Feki, and Prof. Jalel Bouzid, ENIS, Sfax, Tunisia, for their stimulating and beneficial discussion. We extend our gratitude to Prof. Hafed Mejdoub (FSS, Sfax, Tunisia) for the determination of the N-terminal sequence and to Mr Jamil Jaoua, Head of the English Unit at the Sfax Faculty of Science, for having proofread this paper.

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Authors and Affiliations

  1. Unite Enzyme and Bioconversion, Ecole Nationale d’Ingénieurs de Sfax, Route Soukra km 3.5 B.P.W, 3038, Sfax, Tunisia

    Monia Blibech, Raoudha Ellouz Ghorbel, Ines Fakhfakh & Semia Ellouz Chaabouni

  2. Department of Biochemistry, Physiology and Microbiology, Ghent University, K. L. Ledeganckstraat 35, 9000, Ghent, Belgium

    Patricia Ntarima & Katheleen Piens

  3. Laboratoire de Lipolyse Enzymatique, Ecole Nationale d’Ingénieurs de Sfax, Route Soukra km 3.5 B.P.W, 3038, Sfax, Tunisia

    Abir Ben Bacha

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  1. Monia Blibech
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Correspondence to Semia Ellouz Chaabouni.

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Blibech, M., Ghorbel, R.E., Fakhfakh, I. et al. Purification and Characterization of a Low Molecular Weight of β-Mannanase from Penicillium occitanis Pol6. Appl Biochem Biotechnol 160, 1227–1240 (2010). https://doi.org/10.1007/s12010-009-8630-z

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