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Purification and Characterization of Novel α-Amylase from Bacillus subtilis KIBGE HAS

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Abstract

Purification of extracellular α-amylase from Bacillus subtilis KIBGE HAS was carried out by ultrafiltration, ammonium sulfate precipitation and gel filtration chromatography. The enzyme was purified to homogeneity with 96.3-fold purification with specific activity of 13011 U/mg. The molecular weight of purified α-amylase was found to be 56,000 Da by SDS-PAGE. Characteristics of extracellular α-amylase showed that the enzyme had a Km and V max value of 2.68 mg/ml and 1773 U/ml, respectively. The optimum activity was observed at pH 7.5 in 0.1 M phosphate buffer at 50°C. The amino acid composition of the enzyme showed that the enzyme is rich in neutral/non polar amino acids and less in acidic/polar and basic amino acids. The N-terminal protein sequence of 10 residues was found to be as Ser-Ser-Asn-Lys-Leu-Thr-Thr-Ser-Trp-Gly (S-S-N-K-L-T-T-S-W-G). Furthermore, the protein was not N-terminally blocked. The sequence of α-amylase from B. subtilis KIBGE HAS was a novel sequence and showed no homology to other reported α-amylases from Bacillus strain.

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Acknowledgments

The authors would like to thank Dr. Tanzeel Haider Usmani, Director General PCSIR Laboratories Complex, Karachi for kindly providing the lab facilities.

Conflict of interest

The authors have declared that no conflict of interest exists.

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Authors and Affiliations

  1. Pharmaceutical Research Center, PCSIR Laboratories Complex, Karachi, Pakistan

    Saeeda Bano & Muhammad Noman Syed

  2. The Karachi Institute of Biotechnology and Genetic Engineering (KIBGE), University of Karachi, Karachi, 75270, Pakistan

    Shah Ali Ul Qader, Afsheen Aman & Abid Azhar

Authors
  1. Saeeda Bano
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  2. Shah Ali Ul Qader
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  3. Afsheen Aman
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  4. Muhammad Noman Syed
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  5. Abid Azhar
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Correspondence to Shah Ali Ul Qader.

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Bano, S., Qader, S.A.U., Aman, A. et al. Purification and Characterization of Novel α-Amylase from Bacillus subtilis KIBGE HAS. AAPS PharmSciTech 12, 255–261 (2011). https://doi.org/10.1208/s12249-011-9586-1

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  • DOI: https://doi.org/10.1208/s12249-011-9586-1

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