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Molecular Biotechnology

, Volume 58, Issue 1, pp 12–21 | Cite as

Deleting the Ig-Like Domain of Alicyclobacillus acidocaldarius Endoglucanase Cel9A Causes a Simultaneous Increase in the Activity and Stability

  • Fereshteh S. Younesi
  • Mohammad Pazhang
  • Saeed Najavand
  • Parastou Rahimizadeh
  • Mohsen Akbarian
  • Mehdi Mohammadian
  • Khosro Khajeh
Research

Abstract

Endoglucanase Cel9A from Alicyclobacillus acidocaldarius (AaCel9A) is a monomeric enzyme with 537 residues. This enzyme has an Ig-like domain in the N-terminus of the catalytic domain. In this study, the role of the Ig-like domain on the activity, stability, and structural rigidity of AaCel9A and the effect of calcium on enzyme activity and stability were examined by comparing a truncated enzyme with deletion of the Ig-like domain (AaCel9AΔN) to the wild-type enzyme. Our results showed that the deletion of the Ig-like domain increased the catalytic efficiency of the truncated enzyme up to threefold without any significant changes in the K m of the enzyme. Furthermore, pH and temperature optimum for activity were shifted from 6.5 to 7.5 and from 65 to 60 °C, respectively, by deletion of the Ig-like domain. The thermal stability and fluorescence quenching results indicated that the stability and rigidity of the truncated enzyme have been more than that of the wild-type enzyme. Calcium similarly increased the catalytic efficiency of the enzymes (up to 40 %) and remarkably raised the stability of the AaCel9A compared to the AaCel9AΔN. This shows that Ig-like domain has a role in the increase of the enzyme stability by calcium in the wild-type enzyme.

Keywords

Endoglucanase Cel9A Ig-like domain Catalytic efficiency Thermal stability Enzyme rigidity Calcium 

Abbreviations

AaCel9A

Alicyclobacillus acidocaldarius endoglucanase Cel9A

AaCel9AΔN

AaCel9A without Ig-like domain

CBD

carbohydrate binding domain

CMC

Carboxymethylcellulose

DNS

3,5-Dinitrosalicylic acid

GH

Glycoside hydrolase

Ig-like

Immunoglobulin- like

Notes

Acknowledgments

We thank Professor S. Moréra (from Laboratoired’Enzymologie et BiochimieStructurales (LEBS), CNRS, Avenue de la Terrasse, 91198 Gif-sur-Yvette, France) for the gift of pDEST17-AaCel9A. The authors express their gratitude to the research council of Azarbaijan Shahid Madani University for the financial support during the course of this project.

Compliance with Ethical Standards

Conflict of interest

The authors declare that there are no conflicts of interest.

Supplementary material

12033_2015_9900_MOESM1_ESM.docx (136 kb)
Supplementary material 1 (DOCX 135 kb)

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Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Fereshteh S. Younesi
    • 1
  • Mohammad Pazhang
    • 1
  • Saeed Najavand
    • 1
  • Parastou Rahimizadeh
    • 1
  • Mohsen Akbarian
    • 1
  • Mehdi Mohammadian
    • 2
  • Khosro Khajeh
    • 3
  1. 1.Department of Cellular and Molecular Biology, Faculty of ScienceAzarbaijan Shahid Madani UniversityTabrizIran
  2. 2.Department of Nanobiotechnology, Faculty of Biological ScienceTarbiat Modares UniversityTehranIran
  3. 3.Department of Biochemistry, Faculty of Biological ScienceTarbiat Modares UniversityTehranIran

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