Purification, biochemical, and molecular characterization of a novel extracellular thermostable and alkaline α-amylase from Tepidimonas fonticaldi strain HB23.
The present study investigated the purification, biochemical, and molecular characterization of a novel thermostable α-amylase (TfAmy48) from Tepidimonas fonticaldi strain HB23. MALDI-TOF/MS analysis indicated that the purified enzyme is a monomer with a molecular mass of 48,138.10 Da. The results from amino-acid sequence analysis revealed high homology between the 25 NH2-terminal residues of TfAmy48 and those of Gammaproteobacteria α-amylases. The optimum pH and temperature values for α-amylase activity were pH 8 and 80 °C, respectively. Thin-layer chromatography (TLC) analysis showed that the final hydrolyzed products of the enzyme from soluble potato starch were maltopentaose, maltose, and maltotriose, which indicate that TfAmy48 possessed an endo-acting pattern. Compared to Termamyl®300 L, TfAmy48 showed extreme stability and tolerance towards organic solvents and excellent compatibility with some commercial laundry detergents. These proprieties make TfAmy48 enzyme a potential candidate as a cleaning bioadditive in detergent composition. The Tfamy48 gene encoding TfAmy48 was cloned, sequenced, and heterologously-expressed in the extracellular fraction of Escherichia coli strain BL21(DE3)pLysS. The biochemical properties of the extracellular purified recombinant enzyme (rTfAmy48) were similar to those of native one. The highest sequence identity value (97%) was obtained with PsAmy1 α-amylase from Pseudomonas sp. strain KFCC10818, with only 16 amino-acid (aa) residues of difference.
['Allala|Fawzi|F|', 'Bouacem|Khelifa|K|', 'Boucherba|Nawel|N|', 'Azzouz|Zahra|Z|', 'Mechri|Sondes|S|', 'Sahnoun|Mouna|M|', 'Benallaoua|Said|S|', 'Hacene|Hocine|H|', 'Jaouadi|Bassem|B|', 'Bouanane-Darenfed|Amel|A|']
D000595:Amino Acid Sequence D001483:Base Sequence D000073556:Burkholderiales D003001:Cloning, Molecular D004791:Enzyme Inhibitors D004795:Enzyme Stability D005110:Extracellular Space D006863:Hydrogen-Ion Concentration D008670:Metals D008958:Models, Molecular D008970:Molecular Weight D000072417:Protein Domains D017422:Sequence Analysis, DNA D013696:Temperature D000516:alpha-Amylases
Purification α-Amylase Tepidimonas fonticaldi Detergent formulations Heterologous-expression Hydrolytic pattern