Overexpression, purification, and biochemical characterization of the esterase Est0796 from Lactobacillus plantarum WCFS1.
Lactobacillus plantarum is a lactic acid bacterium that is encountered in an extensive range of foods such as fermented dairy products, meat, vegetables, and bakery products. Given the little molecular information available on the lipolytic activity of L. plantarum, the aim of this study was to clone, purify, and biochemically characterize the esterase coded by gene lp_0796 (Est0796). The esterase was cloned in pET28a and purified in two steps, using solid ammonium sulfate and His tag affinity chromatography. The molecular mass of the purified Est0796 was 28.7 kDa (by SDS-PAGE) and 26.6 kDa (by gel filtration chromatography), pointing to a monomeric structure. Est0796 showed maximum activity at pH 8.0 and 35 °C and toward shorter acyl chain lengths (C2-C4). The activity was resistant to organic solvents and cations, suggesting that this esterase may play a role in the fermentation of food products.
Inmaculada
I
Navarro-González
Department of Biochemistry and Molecular Biology-A, University of Murcia, Murcia, Spain. inmaculada.navarro@um.es
Navarro-González
NG
Inmaculada
Sánchez-Ferrer
SF
Alvaro
García-Carmona
GC
Francisco
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D002850:Chromatography, Gel / Q000379:methods; D003001:Cloning, Molecular / Q000379:methods; D003611:Dairy Products / Q000382:microbiology; D004950:Esterases / Q000235:genetics* / Q000378:metabolism; D005285:Fermentation; D005516:Food Microbiology; D006863:Hydrogen-Ion Concentration; D048191:Lactiplantibacillus plantarum / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011994:Recombinant Proteins; D016415:Sequence Alignment
D001426:Bacterial Proteins; D011994:Recombinant Proteins; D004950:Esterases