Characterization of a pH and detergent-tolerant, cold-adapted type I pullulanase from Exiguobacterium sp. SH3.
A pullulanase-encoding gene from psychrotrophic Exiguobacterium sp. SH3 was cloned and expressed in both E. coli and Bacillus subtilis. The functional recombinant enzyme (Pul-SH3) was purified as a His-tagged protein. Pul-SH3 was characterized to be a cold-adapted type I pullulanase with maximum activity at 45 °C. Using fluorescence spectroscopy, the melting temperature of Pul-SH3 was determined to be about 52 °C. The enzyme was able to hydrolyze pullulan, soluble starch, potato starch, and rice flour. It was exceptionally tolerant in the pH range of 4-11, exhibiting maximum activity at pH 8.5 and more than 60% of the activity in the pH range of 5-11. Being a detergent-tolerant pullulanase, Pul-SH3 retained 99, 89, and 54% of its activity at 10% concentration of Triton-X100, Tween 20, and SDS, respectively. The enzyme also exhibited an activity of 80.4 and 93.7% in the presence of two commercial detergents, Rika (7.5% v/v) and Fadisheh (2.5% w/v), respectively. The enzyme was even able to remain active by 54.5 and 85% after 10-day holding with the commercial detergents. Thermal stability of the enzyme could w on silica. Pul-SH3 with several industrially important characteristics seems desirable for cold hydrolysis of starch.
Department of Energy and Environmental Biotechnology, National Institute of Genetic Engineering and Biotechnology (NIGEB), Pajoohesh Blvd., Tehran-Karaj Highway, Tehran, P.O. Box 14155-6343, Iran.|Department of Genetics, Faculty of Biological Science, Tarbiat Modares University, Tehran, P.O. Box 14115-154, Iran.
Sarah
S
Rajaei
Department of Energy and Environmental Biotechnology, National Institute of Genetic Engineering and Biotechnology (NIGEB), Pajoohesh Blvd., Tehran-Karaj Highway, Tehran, P.O. Box 14155-6343, Iran.
Kambiz Akbari
KA
Noghabi
Department of Genetics, Faculty of Biological Science, Tarbiat Modares University, Tehran, P.O. Box 14115-154, Iran.
Majid
M
Sadeghizadeh
Department of Energy and Environmental Biotechnology, National Institute of Genetic Engineering and Biotechnology (NIGEB), Pajoohesh Blvd., Tehran-Karaj Highway, Tehran, P.O. Box 14155-6343, Iran. shahbani@nigeb.ac.ir.
Hossein Shahbani
HS
Zahiri
D000595:Amino Acid Sequence; D001406:Bacillaceae / Q000201:enzymology* / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D003080:Cold Temperature; D003902:Detergents / Q000737:chemistry*; D005798:Genes, Bacterial; D006026:Glycoside Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D011489:Protein Denaturation; D013379:Substrate Specificity
D001426:Bacterial Proteins; D003902:Detergents; D006026:Glycoside Hydrolases; C100181:pullulanase
Cold adapted; Exiguobacterium; Starch; Type I pullulanase