To determine the effect of different temperature on strawberry after harvest, physiological indicator analysis and proteomics analysis were conducted on ripened strawberry (‘Sweet Charlie’) fruit stored at 4 °C, 23 °C, and 37 °C (±2) for 10 or 20 days. Results showed that 4 °C maintained a better visual quality of strawberry, and the weight loss and firmness remained stable within 3 days. Low temperature negatively affected anthocyanin but positively affected soluble sugars. Though anthocyanin content was higher with increasing temperature, anthocyanin synthesis related proteins were downregulated. Higher indole-acetic acid (IAA) content in seeds and lower abscisic acid (ABA) content were found in berry at 4 °C. Antioxidant related proteins were upregulated during storage, showing a significant up-regulation of POD at 4 °C, and AsA-GSH cycle related proteins and heat shock proteins (HSPs) at 37 °C. In addition, overexpressed sugar phosphate/phosphate translocator, 1-aminocyclopropane-1-carboxylate oxidase and aquaporin PIP2-2 had a positive effect in response to low temperature stress for containing higher protopectin content and POD activity.