PMID: 8937850Nov 1, 1996Paper

Improved bacterial expression of the human P form phenolsulfotransferase. Applications to drug metabolism

Drug Metabolism and Disposition : the Biological Fate of Chemicals
A J LewisT Walle

Abstract

Bacterial expression of human phenol phenolsulfotransferase (P-PST) has provided the opportunity to understand better the catalytic properties and biological role of this enzyme. However, as the yield of pure protein from the currently used expression system was low, we subcloned the P-PST c-DNA into pET-15b, a vector containing an oligohistidine domain, for improved expression. The fusion protein, His-P-PST, was isolated from the bacterial cytosol in a single affinity chromatography step, using a Ni2+ agarose column. The yield of His-P-PST from the pET-15b vector was improved 12-fold, compared with P-PST from the original vector. The purity was > 99%, as established by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and densitometry scanning. The enzyme was stable for at least 3 weeks when stored in 20% glycerol at -80 degrees C. A very rapid deterioration of the enzyme during 37 degrees C incubations was effectively prevented by the addition of bovine serum albumin. The sulfonation of several substrates was very similar for His-P-PST and P-PST, with Vmax/KM values (first order rate constants) for the high-affinity substrate p-nitrophenol of 143 +/- 27 and 120 +/- 25 ml min-1 microgram-1 PST [mean +/- SE; not signifi...Continue Reading

Related Concepts

Trending Feeds

COVID-19

Coronaviruses encompass a large family of viruses that cause the common cold as well as more serious diseases, such as the ongoing outbreak of coronavirus disease 2019 (COVID-19; formally known as 2019-nCoV). Coronaviruses can spread from animals to humans; symptoms include fever, cough, shortness of breath, and breathing difficulties; in more severe cases, infection can lead to death. This feed covers recent research on COVID-19.

Blastomycosis

Blastomycosis fungal infections spread through inhaling Blastomyces dermatitidis spores. Discover the latest research on blastomycosis fungal infections here.

Nuclear Pore Complex in ALS/FTD

Alterations in nucleocytoplasmic transport, controlled by the nuclear pore complex, may be involved in the pathomechanism underlying multiple neurodegenerative diseases including Amyotrophic Lateral Sclerosis and Frontotemporal Dementia. Here is the latest research on the nuclear pore complex in ALS and FTD.

Applications of Molecular Barcoding

The concept of molecular barcoding is that each original DNA or RNA molecule is attached to a unique sequence barcode. Sequence reads having different barcodes represent different original molecules, while sequence reads having the same barcode are results of PCR duplication from one original molecule. Discover the latest research on molecular barcoding here.

Chronic Fatigue Syndrome

Chronic fatigue syndrome is a disease characterized by unexplained disabling fatigue; the pathology of which is incompletely understood. Discover the latest research on chronic fatigue syndrome here.

Evolution of Pluripotency

Pluripotency refers to the ability of a cell to develop into three primary germ cell layers of the embryo. This feed focuses on the mechanisms that underlie the evolution of pluripotency. Here is the latest research.

Position Effect Variegation

Position Effect Variagation occurs when a gene is inactivated due to its positioning near heterochromatic regions within a chromosome. Discover the latest research on Position Effect Variagation here.

STING Receptor Agonists

Stimulator of IFN genes (STING) are a group of transmembrane proteins that are involved in the induction of type I interferon that is important in the innate immune response. The stimulation of STING has been an active area of research in the treatment of cancer and infectious diseases. Here is the latest research on STING receptor agonists.

Microbicide

Microbicides are products that can be applied to vaginal or rectal mucosal surfaces with the goal of preventing, or at least significantly reducing, the transmission of sexually transmitted infections. Here is the latest research on microbicides.