Iron(II)- and 2-(oxo)-glutarate-dependent oxygenases catalyze diverse oxidative transformations that are often initiated by abstraction of hydrogen from carbon by iron(IV)-oxo (ferryl) complexes. Control of the relative orientation of the substrate C–H and ferryl Fe–O bonds, primarily by direction of the oxo group into one of two cis-related coordination sites (termed inline and offline), may be generally important for control of the reaction outcome. Neither the ferryl complexes nor their fleeting precursors have been crystallographically characterized, hindering direct experimental validation of the offline hypothesis and elucidation of the means by which the protein might dictate an alternative oxo position. Comparison of high-resolution x-ray crystal structures of the substrate complex, an Fe(II)-peroxysuccinate ferryl precursor, and a vanadium(IV)-oxo mimic of the ferryl intermediate in the L-arginine 3-hydroxylase, VioC, reveals coordinated motions of active site residues that appear to control the intermediate geometries to determine reaction outcome.
SignificanceConversion of ribonucleotides to the 2′-deoxyribonucleotides required for DNA biosynthesis is catalyzed by ribonucleotide reductases (RNRs) via a free-radical mechanism. Known types of RNRs all depend on redox-active transition metals—manganese, iron, or cobalt—for radical initiation. Pathogenic bacteria are challenged by transition metal sequestration and infliction of oxidative stress by their hosts, and the deployment of multiple RNRs with different metal requirements and radical-initiating oxidants is a known bacterial countermeasure. A class I RNR from two bacterial pathogens completely lacks transition metals in its active state and uses a tyrosine-derived dihydroxyphenylalanine radical as its initiator, embodying a novel tactic to combat transition metal- and oxidant-mediated innate immunity and reinforcing bacterial RNRs as potential antibiotic targets.
α-Solanine, a naturally occurring steroidal glycoalkaloid found in nightshade (Solanum nigrum Linn.), was found to inhibit proliferation and induce apoptosis of tumor cells. However, the mechanism involved in suppression of cancer cell metastasis by α-solanine remains unclear. This study investigates the suppression mechanism of α-solanine on motility of the human prostate cancer cell PC-3. Results show that α-solanine reduces the viability of PC-3 cells. When treated with non-toxic doses of α-solanine, cell invasion is markedly suppressed by α-solanine. α-Solanine also significantly elevates epithelial marker E-cadherin expression, while it concomitantly decreases mesenchymal marker vimentin expression, suggesting it suppresses epithelial-mesenchymal transition (EMT). α-Solanine reduces the mRNA level of matrix metalloproteinase-2 (MMP-2), MMP-9 and extracellular OPEN ACCESSMolecules 2014, 19 11897 inducer of matrix metalloproteinase (EMMPRIN), but increases the expression of reversion-inducing cysteine-rich protein with kazal motifs (RECK), and tissue inhibitor of metalloproteinase-1 (TIMP-1) and TIMP-2. Immunoblotting assays indicate α-solanine is effective in suppressing the phosphorylation of phosphatidylinositide-3 kinase (PI3K), Akt and ERK. Moreover, α-solanine downregulates oncogenic microRNA-21 (miR-21) and upregulates tumor suppressor miR-138 expression. Taken together, the results suggest that inhibition of PC-3 cell invasion by α-solanine may be, at least in part, through blocking EMT and MMPs expression. α-Solanine also reduces ERK and PI3K/Akt signaling pathways and regulates expression of miR-21 and miR-138. These findings suggest an attractive therapeutic potential of α-solanine for suppressing invasion of prostate cancer cell.
Most known probes for activity-based protein profiling (ABPP) use electrophilic groups that tag a single type of nucleophilic amino acid to identify cases in which its hyper-reactivity underpins function. Much important biochemistry derives from electrophilic enzyme cofactors, transient intermediates, and labile regulatory modifications, but ABPP probes for such species are underdeveloped. Here, we describe a versatile class of probes for this less charted hemisphere of the proteome. The use of an electron-rich hydrazine as the common chemical modifier enables covalent targeting of multiple, pharmacologically important classes of enzymes bearing diverse organic and inorganic cofactors. Probe attachment occurs by both polar and radicaloid mechanisms, can be blocked by molecules that occupy the active sites, and depends on the proper poise of the active site for turnover. These traits will enable the probes to be used to identify specific inhibitors of individual members of these multiple enzyme classes, making them uniquely versatile among known ABPP probes.
The mammalian target of rapamycin (mTOR) is a serine/threonine protein kinase that belongs to the phosphoinositide-3-kinase (PI3K)-related kinase family. Oncogenic activation of mTOR signaling significantly contributes to the progression of different types of cancers including osteosarcoma (OS; the most common primary malignant tumor of bone). In the present study, we review the association of the mTOR signaling pathway with OS, and the possible effective treatment strategies by targeting this pathway. In the metastatic behavior of OS, one of the most common actionable aberrations was found in the PI3K/Akt/mTOR pathway. Upon phosphorylation, activated mTOR contributes to OS cellular transformation and poor cancer prognosis via downstream effectors such as S6K1, 4EBP1 and eIF4E, which are overexpressed in OS. Targeting the mTOR complex is a significant approach in cancer therapeutic research, and of course, rapamycin is the primary inhibitor of mTOR. Various other chemotherapeutic molecules have also shown potential activity against mTOR. As mTOR is a new promising oncological target and blockade of the mTOR pathway with selective inhibitors has significant potential in OS therapeutic research, the development of the optimal dose, regimen and a rationale for the use of mTOR inhibitors in combination with other anticancer agents may provide a successful treatment strategy for OS.
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