Anti-CASP2 polyclonal antibody

Context: Cancer cells exert differential responses to chemotherapeutics and inhibitors. To the best of our knowledge, a few or no research has been performed until now to determine the effect of EF-24 and RAD001 on MDA-MB-231 breast cancer cells with regard to mRNA expression of apoptotic and anti-apoptotic genes. Aims: In this study, we aimed to investigate the mRNA expression levels of apoptotic (caspase 2 [CASP2], CASP8, and CASP9) and anti-apoptotic (B-cell lymphoma 2 [BCL2] and BCL2-like protein 1 [BCL2L1]) genes after exposure to paclitaxel, EF-24, and RAD001 in MDA-MB-231 cells. Materials and Methods: After treatment, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was used to measure cell viability. mRNA expressions were analyzed using quantitative real-time polymerase chain reaction. Results: Decrease in cell viability ratios was seen in a dose-dependent manner for all chemicals. MDA-MB-231 cells responded slightly different to paclitaxel, EF-24, and RAD001 at the transcriptional level of apoptotic and anti-apoptotic genes. Conclusions: Our results showed that response of these cells to paclitaxel, EF-24, and RAD001 was found different at the transcriptional level of apoptotic and antiapoptotic genes. Therefore, understanding transcriptional changes after these drug exposure may give us a change to figure out more realistic results of the apoptotic pathway inhibition.

Background: In the search for generic expression strategies for mammalian protein families several bacterial expression vectors were examined for their ability to promote high yields of soluble protein. Proteins studied included cell surface receptors (Ephrins and Eph receptors, CD44), kinases (EGFR-cytoplasmic domain, CDK2 and 4), proteases (MMP1, CASP2), signal transduction proteins (GRB2, RAF1, HRAS) and transcription factors (GATA2, Fli1, Trp53, Mdm2, JUN, FOS, MAD, MAX). Over 400 experiments were performed where expression of 30 full-length proteins and protein domains were evaluated with 6 different N-terminal and 8 C-terminal fusion partners. Expression of an additional set of 95 mammalian proteins was also performed to test the conclusions of this study. Results: Several protein features correlated with soluble protein expression yield including molecular weight and the number of contiguous hydrophobic residues and low complexity regions. There was no relationship between successful expression and protein pI, grand average of hydropathicity (GRAVY), or sub-cellular location. Only small globular cytoplasmic proteins with an average molecular weight of 23 kDa did not require a solubility enhancing tag for high level soluble expression. Thioredoxin (Trx) and maltose binding protein (MBP) were the best N-terminal protein fusions to promote soluble expression, but MBP was most effective as a C-terminal fusion. 63 of 95 mammalian proteins expressed at soluble levels of greater than 1 mg/l as N-terminal H10-MBP fusions and those that failed possessed, on average, a higher molecular weight and greater number of contiguous hydrophobic amino acids and low complexity regions. Conclusions: By analysis of the protein features identified here, this study will help predict which mammalian proteins and domains can be successfully expressed in E. coli as soluble product and also which are best targeted for a eukaryotic expression system. In some cases proteins may be truncated to minimise molecular weight and the numbers of contiguous hydrophobic amino acids and low complexity regions to aid soluble expression in E. coli.