Esterase

About: Esterase is a research topic. Over the lifetime, 7622 publications have been published within this topic receiving 168270 citations.

Subcellular localization of rat brain esterases

Abstract: The localization and properties of soluble and bound esterases of subcellular fractions of rat brain have been investigated. Bound esterases were extracted with 1% Triton X-100 and separated by starch gel electrophoresis. By these means a molecular population of isoenzymes was demonstrable that was quantitatively different from the isoenzyme population of the watersoluble esterase activity. The highest specific activity for α-naphthyl acetate hydrolysis was contained in the microsomal fraction and could be extracted by Triton. In contradistinction to whole brain and to other subcellular fractions, microsomes contained a molecular population of esterase isozymes which was qualitatively distinct from that of water extracts in that the fast moving A-type esterases were absent. In addition, there was present a heavy concentration of slow movinig B-esterase. Acetylcholinesterase could also be extracted from this fraction by Triton, migrated with B-esterase and actively hydrolyzed αnaphthyl acetate. Combined el...

Understanding promiscuous amidase activity of an esterase from Bacillus subtilis

Abstract: Water works. Bacillus subtilis esterase BS2 is a promiscuous esterase that shows amidase activity. This amidase activity was shown to depend on a hydrogen-bond network with the substrate amide hydr ...

A novel esterase from a marine mud metagenomic library for biocatalytic synthesis of short-chain flavor esters

Abstract: Marine mud is an abundant and largely unexplored source of enzymes with unique properties that may be useful for industrial and biotechnological purposes. However, since most microbes cannot be cultured in the laboratory, a cultivation-independent metagenomic approach would be advantageous for the identification of novel enzymes. Therefore, with the objective of screening novel lipolytic enzymes, a metagenomic library was constructed using the total genomic DNA extracted from marine mud. Based on functional heterologous expression, 34 clones that showed lipolytic activity were isolated. The five clones with the largest halos were identified, and the corresponding genes were successfully overexpressed in Escherichia coli. Molecular analysis revealed that these encoded proteins showed 48–79 % similarity with other proteins in the GenBank database. Multiple sequence alignment and phylogenetic tree analysis classified these five protein sequences as new members of known families of bacterial lipolytic enzymes. Among them, EST4, which has 316 amino acids with a predicted molecular weight of 33.8 kDa, was further studied in detail due to its strong hydrolytic activity. Characterization of EST4 indicated that it is an alkaline esterase that exhibits highest hydrolytic activity towards p-nitrophenyl butyrate (specific activity: 1389 U mg−1) at 45 °C and pH 8.0. The half-life of EST4 is 55 and 46 h at 40 and 45 °C, respectively, indicating a relatively high thermostability. EST4 also showed remarkable stability in organic solvents, retaining 90 % of its initial activity when incubated for 12 h in the presence of hydrophobic alkanes. Furthermore, EST4 was used as an efficient whole-cell biocatalyst for the synthesis of short-chain flavor esters, showing high conversion rate and good tolerance for high substrate concentrations (up to 3.0 M). These results demonstrate a promising potential for industrial scaling-up to produce short-chain flavor esters at high substrate concentrations in non-aqueous media. This manuscript reports unprecedented alcohol tolerance and conversion of an esterase biocatalyst identified from a marine mud metagenomic library. The high organic solvent tolerance and thermostability of EST4 suggest that it has great potential as a biocatalyst.

[45] Mast cell proteases

Abstract: Publisher Summary This chapter presents a procedure for purification and assay of mast cell proteases. For Purification procedure mast cells are collected from Sprague-Dawley rats by washing the peritoneal cavities with 10 ml of ice-cold phosphate-buffered saline solution at pH 7.2. All subsequent steps are carried out at 4°C, including affinity-adsorption chromatography and adsorption to barium sulfate. During the purification of the mast cell protease, it is important to maintain the solutions at relatively high ionic strength to prevent the enzyme from adsorbing to surfaces. The assay measures chymotrypsin-like esterase activity. The procedure involves mixing of substrate and diluted enzyme appended with the buffer. The absorption change at 256 nm is recorded for about 5 rain. An enzyme unit is defined as an amount of enzyme activity that results in the hydrolysis of 1 μmol of substrate per minute at pH 7.8, 25°C.

Isolation and characterization of a heavy metal-resistant, thermophilic esterase from a Red Sea brine pool.

Abstract: The Red Sea Atlantis II brine pool is an extreme environment that displays multiple harsh conditions such as high temperature, high salinity and high concentrations of multiple, toxic heavy metals. The survival of microbes in such an environment by utilizing resistant enzymes makes them an excellent source of extremophilic enzymes. We constructed a fosmid metagenomic library using DNA isolated from the deepest and most secluded layer of this pool. We report the isolation and biochemical characterization of an unusual esterase: EstATII. EstATII is thermophilic (optimum temperature, 65°C), halotolerant (maintains its activity in up to 4.5 M NaCl) and maintains at least 60% of its activity in the presence of a wide spectrum of heavy metals. The combination of biochemical characteristics of the Red Sea Atlantis II brine pool esterase, i.e., halotolerance, thermophilicity and resistance to heavy metals, makes it a potentially useful biocatalyst.