Novel inhibitors of As(III) S-adenosylmethionine methyltransferase (AS3MT) identified by virtual screening.

Due to increased interest in As(III) S-adenosylmethionine methyltransferase (AS3MT), a search for chemical probes that can help elucidate function was initiated. A homology model was built based on related enzymes, and virtual screening produced 426 potential hits. Evaluation of these compounds in a functional enzymatic assay revealed several modest inhibitors including an O-substituted 2-amino-3-cyano indole scaffold.

A high-throughput screening-compatible homogeneous time-resolved fluorescence assay measuring the glycohydrolase activity of human poly(ADP-ribose) glycohydrolase.

Poly(ADP-ribose) (PAR) polymers are transient post-translational modifications, and their formation is catalyzed by poly(ADP-ribose) polymerase (PARP) enzymes. A number of PARP inhibitors are in advanced clinical development for BRCA-mutated breast cancer, and olaparib has recently been approved for BRCA-mutant ovarian cancer; however, there has already been evidence of developed resistance mechanisms. Poly(ADP-ribose) glycohydrolase (PARG) catalyzes the hydrolysis of the endo- and exo-glycosidic bonds within the PAR polymers.

Quantitative investigation of the affinity of human respiratory syncytial virus phosphoprotein C-terminus binding to nucleocapsid protein.

Background: There are no approved small molecule drug therapies for human respiratory syncytial virus (hRSV), a cause of morbidity and mortality in at-risk newborns, the immunocompromised, and the elderly. We have investigated as a potential novel hRSV drug target the protein-protein interaction between the C-terminus of the viral phosphoprotein (P) and the viral nucleocapsid protein (N), components of the ribonucleoprotein complex that contains, replicates, and transcribes the viral RNA genome. Earlier work by others established that the 9 C-terminal residues of P are necessary and sufficient for binding to N.

Enzymatic characterisation of USP7 deubiquitinating activity and inhibition.

USP7 (HAUSP) is a deubiquitinating enzyme, which plays a crucial role in regulating the levels of the p53 tumour suppressor protein, through its ability to prevent the proteasomal degradation of the Ubiquitin ligase for p53, Hdm2. Supporting evidence suggests that an inhibitor of USP7 would act to abrogate the action of Hdm2, and thereby elevate levels of the p53 protein, with associated therapeutic benefits in cancer and potentially other diseases. In this article, we describe the characterisation of differential enzyme activity of both the full length and putative catalytic domain of human USP7 expressed in both bacterial and insect cell expression systems.

Improvements to the throughput of recombinant protein expression in the baculovirus/insect cell system.

Recombinant baculoviruses have proved to be a very useful means to express many proteins over the last 20 years. Since their introduction, there have been a number of significant improvements that have simplified and speeded up the construction of baculoviruses. One of the most commonly used methods relies upon recombination with the baculovirus genome maintained in Escherichia coli.