Chemical biology tools to study Deubiquitinases and Ubl proteases

The reversible attachment of ubiquitin (Ub) and ubiquitin like modifiers (Ubls) to proteins are crucial post-translational modifications (PTMs)for many cellular processes. Not only do cells possess hundreds of ligases to mediate substrate specific modification with Ub and Ubls, but they also have a repertoire of more than 100 dedicated enzymes for the specific removal of ubiquitin (Deubiquitinases or DUBs) and Ubl modifications (Ubl-specific proteases or ULPs). Over the past two decades, there has been significant progress in our understanding of how DUBs and ULPs function ata molecular level and many novel DUBs and ULPs, including several new DUB classes, have been identified. Here, the development of chemical tools that can bind and trap active DUBs has played a key role. Since the introduction of the first activity-based probe for DUBs in 1986, several innovations have led to the development of more sophisticated tools to study DUBs and ULPs. In this review we discuss how chemical biology has led to the development of activity-based probes and substrates that have been invaluable to the study of DUBs and ULPs. We summarise our currently available toolbox, highlight the main achievements and give an outlook of how these tools may be applied to gain a better understanding of the regulatory mechanisms of DUBs and ULPs

MOLECULAR DOCKING, DESIGN, SYNTHESIS, IN VITRO ANTIOXIDANT AND ANTI-INFLAMMATORY EVALUATIONS OF NEW ISOQUINOLINE DERIVATIVES

Objectives: To design and synthesis N-substituted (E)-4-arylidene-isoquinoline-1,3-dione derivatives as anti-inflammatory and antioxidant drug moieties using molecular docking as a tool.Methods: The structure of compounds (5a-h) was elucidated by means of FT-IR, GC-MS and NMR spectroscopy. Molecular docking was carried out to clarify the molecular aspects of the observed COX-inhibitory activities of the investigated compounds. DPPH radical scavenging analysis method was used to determine antioxidant activity and in-vitro anti-Inflammatory activity was conducted by Human Red Blood Cell (HRBC) membrane stabilization method utilizing Diclofenac sodium as standard.Results: Isoquinoline (N-substituted (E)-4-arylidene-isoquinoline-1,3-dione) derivatives were achieved using oxalic acid as the catalyst, by aldol condensation of the corresponding aldehydes and the corresponding N-substituted homophthalimides with a maximum yield of 92%. Ligand efficiency (LE) consequences being a clear indication that the action potential of the compounds 5e (-0.72) and 5d (-0.64) is high when compare with the standard (-0.63) for COX-1. While for COX-2, compounds 5e (-0.81) 5d (-0.79) and 5h (-0.98) were shown a remarkable ligand efficiency than the standard (-0.65). Anti-inflammatory and antioxidant studies on the compounds 5h<5d<5g was found best activity results.Conclusion: From our overall studies, it was understood that the activities of both in silico and in vitro anti-inflammatory results are coincide together. The p-values were significant for all the compounds 5(a-h) in both COX-1 and COX-2 activities which indicate that all the compounds have ‘competency' towards druggability for both anti-inflammatory and antioxidant, especially the compounds 5h<5d<5g<5e can be suggested for in vivo.Â

Low salinity cyclic water injection for enhanced oil recovery in Alaska North Slope

Thesis (M.S.) University of Alaska Fairbanks, 2009"Properties and flow pattern of injected water have an impact on properties like rock wettability and oil saturation. Researchers have observed increased oil recovery with low salinity brines and reduced water production with cyclic injection. Low salinity cyclic water injection is an interesting combination to be evaluated for further implementation. Two-phase water-oil flow experiments were conducted on cleaned and oil-aged sandstone cores in a core holder apparatus. At connate water saturation, modified Amott-Harvey tests were performed to study wettability. Cyclic waterfloods were conducted to recover oil. Residual oil saturation (Sor) was calculated after every step. The experiments were repeated with reconstituted brines of different salinity and Alaska North Slope (ANS) lake water. The effect of low salinity waterfloods and oil-aging on wettability alteration was studied. The results were compared with available data from conventional floods performed on the same cores. Cyclic floods were also tested for different pulse intervals. Conventional waterflooding was conducted on recombined oil-saturated cores at reservoir conditions. Faster reduction in Sor and additional oil recovery was observed consistently with low salinity cyclic injection. Oil-aging reduced water wetness of cores. Subsequent low salinity floods restored the water wetness marginally. Shorter pulses yielded better results than longer intervals"--Leaf iiiArctic Energy Technology Development Laboratory, U.S. Department of Energ

OTU deubiquitinases reveal mechanisms of linkage specificity and enable ubiquitin chain restriction analysis

Sixteen ovarian tumor (OTU) family deubiquitinases (DUBs) exist in humans, and most members regulate cell-signaling cascades. Several OTU DUBs were reported to be ubiquitin (Ub) chain linkage specific, but comprehensive analyses are missing, and the underlying mechanisms of linkage specificity are unclear. Using Ub chains of all eight linkage types, we reveal that most human OTU enzymes are linkage specific, preferring one, two, or a defined subset of linkage types, including unstudied atypical Ub chains. Biochemical analysis and five crystal structures of OTU DUBs with or without Ub substrates reveal four mechanisms of linkage specificity. Additional Ub-binding domains, the ubiquitinated sequence in the substrate, and defined S1’ and S2 Ub-binding sites on the OTU domain enable OTU DUBs to distinguish linkage types. We introduce Ub chain restriction analysis, in which OTU DUBs are used as restriction enzymes to reveal linkage type and the relative abundance of Ub chains on substrates