Deacylation Mechanism Of Long-Chain Fatty Acid Modified Or Succinylated Proteins Catalyzed By SIRT3 Or SIRT5

Sirtuins play important roles in many biological processes and age-related diseases. Apart from the NAD+-dependent deacetylase activity, they also possess other activities such as desuccinylation or deacylation of long-chain fatty acyl lysine. To investigate the deacylation mechanism of long-chain fatty acyl lysine by sirtuins, we solved the crystal structures of SIRT3 in complex with a H3K9 myristoylated or palmitoylated peptide. We found that the myristoyl and palmitoyl may bind at different spaces in the C-site of SIRT3 and the C-site could change its conformation in order to interact with the long-chain acyl substrates. These results could help us design modulators of sirtuins, and also elucidate the deacylation mechanism of long-chain acyl lysine by sirtuins.Sirtuins inhibitor may serve as probe to study the biological functions of sirtuins and have therapeutic potential in the age-related diseases. So far, a lot of inhibitors were developed targeting to A, B or C-site of sirtuins. To aid the design of SIRT5 inhibitor, here we solved the crystal structure of SIRT5 in complex with a coumarin-labelled succinyl peptide(ac-LGKsuc-AMC). By comparing the structures of SIRT5/ac-LGKsuc-AMC and SIRT5/H3K9suc(a succinyl H3K9 peptide), we found that the steric hindrance of the coumarin(i.e., AMC) forced the 250-257 residues of SIRT5 to have significant conformational changes. As the result, the residues such as Ser251, Ser252, Val253 approach to the Rossmann fold domain and the gap between the zinc binding domain and the Rossmann fold domain was enlarged. The crystal structure of “SIRT5/ac-LGKsuc-AMC” would help us design new modulator for SIRT5. Meanwhile, we also showed that piceatannol, a resveratrol derivative, is a SIRT5 inhibitor. Based on the results of thermal shift assays and CD experiments, we proposed that piceatannol binds to SIRT5 and constrains its conformational change. Since SIRT5 undergoes a series of conformational changes during desuccinylation, the constrained conformation is not beneficial for the desuccinylation and thus piceatannol inhibits the activity of SIRT5. To elucidate the inhibition mechanism of piceatannol, we need to further determine the binding mode of piceatannol in SIRT5.