Intermittent fasting, which can effectively reduce obesity and improve the related metabolic syndrome has become an exciting research area in recent years. Adipose tissue is pivotal in regulating the metabolism and determining the occurrence of obesity. In the current study, we aimed to investigate the effects of acute fasting (AF) on fat tissue. Mice were subjected to AF for 36 h, receiving normal chow (low-fat diet [LFD]) or a high-fat diet (HFD), with free ad libitum access to drinking water, and those fed on free-diet counterparts without fasting serveding as controls. We found that AF obviously reshaped the morphology of fat tissue (WAT) and promoted the beiging of white adipose tissue in both LFD- and HFD-fed mice. AF principally improved the lipid metabolism, and increased the M2- polarization of macrophages in WAT white fat tissue of HFD-fed mice. Interestingly, we found that AF dramatically upregulated Sirt5 expression levels and fat tissue succinylation, suggesting that AF-induced beneficial effects on fat might occur via the regulation of Sirt5 levels and altered succinylation in fatty tissues. Our study clearly showed the remodeling function of adipose tissue during AF; in terms of mechanism, the regulation of succinylation levels by AF might provide new insights into the mechanism(s) underlying the improvement in fat metabolism by energy restriction.