The use of conformationally restricted phospholipids 1 and 2 has been employed to understand the conformational preference of phospholipase A2 (PLA2) for substrate phospholipids. Inhibition of porcine pancreatic PLA2 with 1 and 2 indicated a two-to fivefold preference for the distal isomer 2 over the proximal isomer 1. Based upon these studies, both side-chains of the substrate phospholipid appear to occupy the lipid binding domains near the active site with the side-chains further apart most preferred by PLA2.
pancreatic PL42
Key w& PL42 substrate conformation, odeling, active-site inhibitors, phospholipid side chain orientation, Phospholipase A2 (PLA2) is a hydrolase responsible for the release of fatty acids from the two position of membrane phospholipids [l]. In particular, the liberation of arachidonic acid, a mediator of inflammation, is regulated by PLA2 [ 1-31. The ability to specifically inhibit PLA2 has been the focus of several labs for the potential discovery of anti-inflammatories.
To further understand the mode of substrate-PLA2 binding, efforts have been directed at using phospholipids as probes. In earlier work by deHaas and coworkers [7], the minimum substrate requirement for PLA2 was shown to be the acylester glycolphosphorylcholine. This discovery led to the preparation of a variety of alkylphosphorylcholines [8-121, which were shown to protect the active site Hisa from alkylation by para-bromophenacyl bromide, a histidine alkylating reagent. Extending this observation to the triglyceride phosphorylcholines [ 13-18] demonstrated the relative importance of the upper side chain for active site phospholipid binding [ 17,181. Typically, greater PLA2 inhibition was observed when the upper side-chain length was extended. These results suggested the upper side-chain orients the triglyceride to afford optimum substrate binding to PLA2. This is consistent with the lower K, associated with triglyceride phospholipid over the glycol phospholipid aimed at the preparation of potential transition-state analogs have resulted in the preparation of glycol and triglyceride phosphorylcholines with difluoroketone substitution of cleavage site.