Purification and characterization of a thermostable esterase from the moderate thermophile Bacillus circulans

Abstract

The thermostable esterase from the moderate thermophile Bacillus circulans was purified to homogeneity using a four-step procedure. Esterase activity was associated with a protein of molecular mass 95 kDa, composed of three identical subunits of 30 kDa. The esterase activity was thermostable with a maximum activity at 55 degrees C using initial rate assay. The half-inactivation temperature was 71 degrees C after a 1-h treatment, which compared favorably to that of other enzymes. Activity at temperatures of 30-37 degrees C was high (about half of maximum), making this new enzyme very attractive for applications in this moderate temperature range. The esterase also showed high activity at a rather alkaline pH (higher than 10). The specificity pattern showed a marked specificity for mid-chain-length fatty acids (3-8 carbon atoms), which classified the enzyme as a carboxylesterase.

Thermostable Esterase from a Thermoacidophilic Archaeon: Purification and Characterization for Enzymatic Resolution of a Chiral Compound

“…11,23,24) Although this categorization has limitations due to overlapping substrate specificities or inhibitor patterns, this classification is generally used to characterize the biochemical properties of esterases. Phenylmethylsulfonyl fluroride (PMSF) and diethyl p-nitrophenyl phosphate (paraoxon) inhibited Est3 activity in the above experiment.…”

Biochemical properties and three-dimensional structures of two extracellular lipolytic enzymes from Bacillus subtilis

“…A number of publications describe the cloning, expression, purification and biochemical characterization of lipases from Bacillus species ( [9][10][11][12][22][23][24][25][26][27][28][29][30][31][32][33][34][35][36][37][38][39]). More recently, the first 3D structure of a Bacillus lipase was solved [41].…”

Indication for a New Lipolytic Enzyme Family: Isolation and Characterization of Two Esterases from a Metagenomic Library

“…Being stable without a significant loss of activity in an acidic as well as in an alkaline milieu sets EstGK1 and EstZ3 apart from most esterases. Esterases are often active in an alkaline milieu [Gupta et al, 2004] and there are few examples of lipolytic enzymes acting in a wider range of pH [Elend et al, 2006;Kademi et al, 2000;Kulkarni and Gadre, 2002;Rusnak et al, 2005], but only the lipase from Pseudomonas fluorescens NS2W [Kulkarni and Gadre, 2002] exhibited a comparable pH spectrum to EstZ3. Both enzymes did not show special features concerning temperature stability.…”