Molecular Study On The Uncoupling Protein-1 Of Tree Shrew

The article is about the cold adaptative characters of the UCP1 on the molecular level during the cold exposure . The Tree Shrew, which was selected as the experimental animal in our research, is he representative tropic and sub-tropic small mammal distributed in the south of China . Our main work is to purify the UCP1 and analysis its sequences.This thesis is consists of three parts:1. The purification of UCP-l(Uncoupling protein -1) from BAT(brown adipose tissue) of the tree shrew.2. The preparation of antisersum of rabbit anti -UCP1 from brown adipose tissue in the tree shrew and enzyme linked immunosorbent assays.3. Analysis by the mean of high performance liquit chromatography .(HPLC). The main results and conclusions are show as follow:1. On the basis of the method of purification for UCP-1 in rodent by Lin et al. The UCP1 from adipose tissue in tree shrew was isolated and purified successfully. The MW of UCP-1 was about 33.142kDa, which is consisted of 306 amino acids. We measured the physical effect of UCP-1 of the BAT under the coldexposure. We found it increased abviously about 7 days cold acclimation. And after 28 day in the cold acclimation (4 ) , the content of the UCP-1 in the mitochondrion, the weight of the BAT and the total content of the protein in the mitochondrion began to smooth. So we conclude that cold exposure is an efficient factor which affect the content of UCP-1 in small mammal animal.2. The antiserum of rabbit anti-UCP1 from brown adipose tissue of tree shrew was prepared successfully . And the enzyme linked immunsorbent assays was established too. In our experiment, we select 0, 0.1, 0.8, 1- 3, 7, 14, 21, 28 days as our gradient by this means . We found the content of the UCP1 and the time of the cold exposure have the positive relation . The content of UCP1 keep stability after 7 days . And between 21 days and 28 days, it was liable to smooth. At the same time , we can conclude from our result: UCP1 and the rabbit anti-UCPl of tree shrew have a single tide-site . The content of rabbit anti-UCPl was about 1000-1000, which was the better content. We know now , UCP1 only can be found in the BAT .UCP2 exist in the tissue all of the body. But UCP3 only exist in BAT and muscle.3. From the map of the HPLC(high performance liquit chromatography), we can know that the thing we get from experiment maybe has 2 kinds of component o The characteristic of them were similar. The have the absorb peak are similar , which was about 218 and 271. Otherwise , the sequences of amino acids of UCP1 and UCP3 have the same sequences about 71 % , and also have the similar MW. UCP1 and UCP3 both the inner protein-carriers in the mitochondrion . Though they both can do the carry work of H+ , but the mechanism are different, UCP1 can uncouple the correlation between theoxidation phophorylation and the respiratory chain or the flux of H+. But, UCP3 only form a protein transporter,which only affect the base metabolism . So in the cold acclimated time, the content of UCP1 increased, but not UCP3. Which is the molecular mechanism of the adaptive thermogenesis.Conclusion:In the course of cold acclimation, the main way of the tree shrew to keep the temperature is based on the UCP1(Uncoupling Protein-1) of the BAT. Although, UCP3 exist in BAT, but is has no the same function , UCP3 also exist in muscle . It has the effect on the base metablism. UCP2 has the extensive display, which maybe regulate the temperature of the special parts.