Engineering of the α‐amylase from Geobacillus stearothermophilus US100 for detergent incorporation

Abstract

AmyUS100DeltaIG is a variant of the most thermoactive and thermostable maltohexaose forming alpha-amylase produced by Geobacillus stearothermophilus sp.US100. This enzyme which was designed to improve the thermostability of the wild-type enzyme has acquired a very high resistance to chelator agents. According to modeling structural studies and with the aim of enhancing its resistance towards chemical oxidation, a mutant (AmyUS100DeltaIG/M197A) was created by substituting methionine 197 to alanine. The catalytic proprieties of the resulting mutant show alterations in the specific activity and the profile of starch hydrolysis. Interestingly, AmyUS100DeltaIG/M197A displayed the highest resistance to oxidation compared to the AmyUS100DeltaIG and to Termamyl300, the well-known commercial amylase used in detergent. Further, performance of the engineered alpha-amylase was estimated in the presence of commonly used detergent compounds and a wide range of commercial detergent (liquid and solid). These studies indicated a high compatibility and performance of AmyUS100DeltaIG/M197A, suggesting its potential application in detergent industry.

Fusion of an Oligopeptide to the N Terminus of an Alkaline α-Amylase from Alkalimonas amylolytica Simultaneously Improves the Enzyme's Catalytic Efficiency, Thermal Stability, and Resistance to Oxidation

“…Oxidation of the methionine residue situated in the cavity of the AmyK active site has been shown to decrease its activity or even inactivate the enzyme (11,33). The model structure of AmyK from Bacillus sp.…”

“…The compatibility of AmyK-p1 with the washing powders may be the result of ionic interactions and hydrophobic or hydrophilic interactions between the detergents and the fusion peptide (11). This might be explained by the fact that for liquid or laundry detergents, the physical isolation of enzymes was difficult and the presence of solvent (water) amplified the detrimental effects of surfactants due to the numerous surfactants in those detergents (11,35).…”

“…This might be explained by the fact that for liquid or laundry detergents, the physical isolation of enzymes was difficult and the presence of solvent (water) amplified the detrimental effects of surfactants due to the numerous surfactants in those detergents (11,35). In addition, AmyK-p1 activity, which is optimal at alkaline pH, was greatly decreased in the presence of liquid detergents with acidic pH (5.5 to 8.0) compared to solid washing powders with pH ranging from 9.0 to 11.0.…”

“…This application requires high catalytic efficiency, high alkaline stability, and high oxidative stability (11). Although site-directed mutagenesis has been used to improve the oxidative stability of ␣-amylases, this was accompanied by decreased catalytic efficiency, alkaline stability, and thermal stability (4,5).…”

Improved thermostable α-amylase activity of Bacillus amyloliquefaciens by low-energy ion implantation

“…In the present study, we used low-energy N + ion implantation to mutate the B. amyloliquefaciens strain RL, an important microbe (with roles in many aspects of industry, agriculture, environmental reclamation, pest control, etc.) that secretes a thermostable α-amylase, and screened for mutants with increased starch degradation activity (Declerck et al, 2000;Kim et al, 2003;Du et al, 2006;Khemakhem et al, 2009). We found that, compared to the parental strain, the mutant strain RL-1 excreted a highly thermostable α-amylase with 57.1% increased activity (58.51 U/mL).…”