We determined the crystal structure of the full-length cAMP phosphodiesterase Rv0805 from Mycobacterium tuberculosis. Furthermore we showed, that localization of Rv0805 to the bacterial cell wall is dependent on its C terminus, and expression of either wild type or mutationally inactivated Rv0805 in M. smegmatis alters cell permeability to hydrophobic cytotoxic compounds. Rv0805 may therefore play a key role in the pathogenicity of mycobacteria, not only by hydrolyzing bacterial cAMP, but also by moonlighting as a protein that can alter cell wall functioning.
We studied the effect of phenobarbital on transcription in livers of wild type, CAR-/-, PXR-/- and CAR/PXR-/- knockout mice. In primary cultures of hepatocytes isolated from CAR/PXR -/- knockout mice, it increased HNF-4alpha levels. We provided evidence that it directly induces transcription of the PPARalpha gene via its HNF-4alpha response element, and indirectly by lack of inhibitory crosstalk of AMPK, CAR and PXR with HNF-4alpha. Our results provide further insight into CAR and PXR independent effects of phenobarbital and the crosstalk between different nuclear receptor signaling pathways.
In K5 and K14 mutant (EBS) keratinocytes we found many genes associated with the cytoskeleton having altered expression levels; in particular cell junction components are down-regulated. That this is due to the expression of the mutant keratins, and not to other genetic variables, is supported by experiments on isogenic cells we generated from wild type keratinocytes transfected with the same K5 and 14 mutations. These findings help explain other aspects of EBS-associated pathology. The weakened cell junctions may be also contributing to the reported increased risk of BCC in EBS patients.
Functional polymorphism 68G)C (Cys23Ser) and promoter polymorphism -995G)A of serotonin receptor 2C (HTR2C) have already been investigated, but no association with suicide was found. However, we observed significant association between female suicide victims and polymorphism 68G)C. The significance remained when we combined alleles of female and male populations. An excess of GG genotype and allele G was observed. No statistically important differences were present when only males were analyzed. Haplotype analysis on female population showed marginal association of haplotype G-C with suicide.
Pegylation is considered one of the most efficient methodologies for half-life extension of biopharmaceutical drugs that also improves physicochemical and biological characteristics of proteins. To explore the impact of polyethylene glycol (PEG) size on 'in vitro' potency, a series of well-defined conjugates of interferon a-2b (IFN) were prepared with PEGs of different lengths and shapes. Beside of the expected impact of the increased molecular size, the studies revealed incredibly high correlation between 'in vitro' potency and chromatographic behavior on the cation exchange chromatography.