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A New Member of Family 11 Polysaccharide Lyase, Rhamnogalacturonan Lyase (CtRGLf) from Clostridium thermocellum

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Abstract

A thermostable, alkaline rhamnogalacturonan lyase (RG lyase) CtRGLf, of family 11 polysaccharide lyase from Clostridium thermocellum was cloned, expressed, purified and biochemically characterised. Both, the full-length CtRGLf (80 kDa) protein and its truncated derivative CtRGL (63.9 kDa) were expressed as soluble proteins and displayed maximum activity against rhamnogalacturonan I (RG I). CtRGLf showed maximum activity at 70 °C, while CtRGL at 60 °C. Both enzymes showed maximum activity at pH 8.5. CtRGLf and CtRGL do not show higher activity on substrates with high β-d-galactopyranose (d-Galp) substitution, this catalytic property deviates from that of some earlier characterised RG lyases which prefer substrates with high d-Galp substitution. The enzyme activity of CtRGLf and CtRGL was enhanced by 1.5 and 1.3 fold, respectively, in the presence of 3 mM of Ca2+ ions. The TLC analysis of the degraded products of RG I, released by the action of CtRGLf and CtRGL revealed the production of RG oligosaccharides as major products confirming their endolytic activity.

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Acknowledgments

The authors thank Department of Science and Technology, Ministry of Science and Technology, New Delhi, Govt. of India and Fundação para a Ciência e a Tecnologia, Portugal for Indo-Portugal joint project grant (INT/Portugal/P-14/2013). Fellowship provided by Ministry of Human Resource Development, Govt. of India to AD is gratefully acknowledged.

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Authors and Affiliations

  1. Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Guwahati, 781039, Assam, India

    Arun Dhillon & Arun Goyal

  2. CIISA-Faculdade de Medicina Veterinária, Universidade de Lisboa, Av. da Universidade de Técnica, 1300-477, Lisbon, Portugal

    Vania O. Fernandes, Fernando M. V. Dias, José A. M. Prates, Luis M. A. Ferreira, Carlos M. G. A. Fontes & M. S. J. Centeno

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  1. Arun Dhillon
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Correspondence to Arun Goyal.

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Dhillon, A., Fernandes, V.O., Dias, F.M.V. et al. A New Member of Family 11 Polysaccharide Lyase, Rhamnogalacturonan Lyase (CtRGLf) from Clostridium thermocellum . Mol Biotechnol 58, 232–240 (2016). https://doi.org/10.1007/s12033-016-9921-6

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