Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.5.1.18 (glutathione S-transferase)
 22,582 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Mouse PNAS-4 (mPNAS-4) has 96% identity with human PNAS-4 (hPNAS-4) in primary sequence and has been reported to be involved in the apoptotic response to DNA damage. However, there have been no studies reported of the biological functions of mPNAS-4. In studies conducted by our group (unpublished data), it was interesting to note that overexpression of mPNAS-4 promoted apoptotic death in Lewis lung carcinoma cells (LL2) and colon carcinoma cells (CT26) of mice both in vitro and in vivo. In our studies, mPNAS-4 was cloned into the pGEX-6P-1 vector with GST tag at N-terminal in Escherichia coli strain BL21(DE3). The soluble and insoluble expression of recombinant protein mPNAS-4 (rmPNAS-4) was temperature-dependent. The majority of rmPNAS-4 was insoluble at 37 degrees C, while it was almost exclusively expressed in soluble form at 20 degrees C. The soluble rmPNAS-4 was purified by one-step affinity purification, using a glutathione Sepharose 4B column. The rmPNAS-4 protein was further identified by electrospray ionization-mass spectrometry analysis. The search parameters of the parent and fragment mass error tolerance were set at 0.1 and 0.05 kDa, respectively, and the sequence coverage of search result was 28%. The purified rmPNAS-4 was further used as immunogen to raise polyclonal antibodies in New Zealand white rabbit, which were suitable to detect both the recombinant and the endogenous mPNAS-4 in mouse brain tissue and LL2 cells after immunoblotting and/or immunostaining. The purified rmPNAS-4 and our prepared anti-mPNAS-4 polyclonal antibodies may provide useful tools for future biological function studies for mPNAS.
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PMID:Prokaryotic expression of a novel mouse pro-apoptosis protein PNAS-4 and application of its polyclonal antibodies. 1862 94

Recently, hPNAS-4 [human PNAS-4; also known as C1orf121 (Genbank accession number NP-057160)] was reported to be a novel pro-apoptotic protein in mammalian cells. However, at present there is little information about hPNAS-4 and its molecular mechanisms. In our present studies, the hPNAS-4 gene was first cloned into the pGEX-6p-1 vector with a GST (glutathione transferase) tag to express in soluble form in Escherichia coli induced with 0.5 mM IPTG (isopropyl beta-D-thiogalactoside) at 25 degrees C, and the recombinant hPNAS-4 was purified to near homogeneity with 96% purity by affinity chromatography and anion-exchange chromatography. The purified hPNAS-4 protein was further identified by liquid chromatography-ESI (electrospray ionization)-MS analysis and used to raise polyclonal antibodies that could specifically recognize the hPNAS-4 protein. In addition, hPNAS-4 is localized to the cytoplasm and the perinuclear compartment. Furthermore, the overexpression of hPNAS-4 in human A549 lung cancer cells resulted in decrease of cell viability via apoptosis. This study represented an important step to investigate the characterization for the new pro-apoptosis protein hPNAS-4, which should aid the discovery of new drug targets for the development of effective therapeutic approaches to cancer in the future.
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PMID:Prokaryotic expression, purification and characterization of a novel pro-apoptosis protein hPNAS-4. 1991 10