Phospholipase
A2, group XVI (PLA2G16) is a thiol hydrolase from
the HRASLS family that regulates lipolysis in adipose tissue and has
been identified as a host factor enabling the cellular entry of picornaviruses.
Chemical tools are essential to visualize and control PLA2G16 activity,
but they have not been reported to date. Here, we show that MB064,
which is a fluorescent lipase probe, also labels recombinant and endogenously
expressed PLA2G16. Competitive activity-based protein profiling (ABPP)
using MB064 enabled the discovery of α-ketoamides as the first
selective PLA2G16 inhibitors. LEI110 was identified as a potent PLA2G16
inhibitor (Ki = 20 nM) that reduces cellular
arachidonic acid levels and oleic acid-induced lipolysis in human
HepG2 cells. Gel-based ABPP and chemical proteomics showed that LEI110
is a selective pan-inhibitor of the HRASLS family of thiol hydrolases
(i.e., PLA2G16, HRASLS2, RARRES3 and iNAT). Molecular dynamic simulations
of LEI110 in the reported crystal structure of PLA2G16 provided insight
in the potential ligand–protein interactions to explain its
binding mode. In conclusion, we have developed the first selective
inhibitor that can be used to study the cellular role of PLA2G16.
The biomass holdup and characteristics of the anareobic fluidised bed system for methane recovery from liquid wastes was examined at COD loadings of between 5.8 to 108 kg m−3 day,−1 hydraulic retention times of between 0.45 to 8h, and feed COD concentrations of between 480 to 9000 mg dm−3. Under these operating conditions, the equilibrium biomass holdups increased with increasing COD loadings and varied from 15000 to 32000 mg VSS dm−3 The distribution of biomass holdup and biofilm thickness in the reactor was relatively uniform, because of the completely mixed conditions maintained and the continuous sloughing of biofilms induced by the effervescence caused by rising methane bubbles. This continuous biofilm sloughing process also eliminated the need for intentional sludge wasting and consequently, the resulting sludge retention time in the reactor decreased with increasing COD loadings. The ability of the anaerobic fluidized bed system to retain a high biomass holdup was clearly demonstrated. As a result this system is ideal for being employed as a high‐rate system for methane recovery from liquid wastes, even at low feed COD concentrations.
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