Mitochondrial Membrane Gap Protein Interaction Network Analysis

Mitochondrion is an organelle that universally exist in eukaryotic cells, is the main place of cell oxidative phosphorylation and adenosine triphosphate (ATP) generation, supplies energy for complex cell biology function. Mitochondrion contains outer membrane, intermembrane space, inner membrane and cristae. Intermembrane space is a 6-8nm space between the outer membrane and the inner membrane to connect cristae with cytosol. The proteins of the intermembrane space participate in electron transport and energy metabolism, material transfer, metabolism, cell proliferation and apoptosis, and so on. There are 29 types proteins have been clearly identified. So few proteins have been involved in so many important cell biology progress, which suggested there should be many connections among these proteins. It is very important to explain these protein function and mechanism by investigating their interactions.We previously used yeast two hybridization combined with aim protein interaction mating method to investigate 41 proteins including 20 intermembrane space proteins, 19 inner/outer membrane proteins that have exposure domain in intermembrane space and 2 cristae proteins which encoded by nuclear gene. We found 49 pairs of non-redundant interactions related to 36 proteins and established the intermembrane space protein interaction network integrated with LCI network. We analyzed the network and validated some interactions of the network with Co-IP and GST-pull down. The investigation is very important to better understand the function establishment of proteomics and the biological characters of intermembrane space proteins. We found 9 proteins located on intermembrane space and 4 inner membrane proteins with exposure domain in the intermembrane space, with these 54 aim proteins including these 13 new proteins we complete the interaction network which can be better explain the protein function and mechanism.We obtained 110 pairs of rotary verification positive interactions related to 75 proteins matched with 31 aim proteins from human liver and brain cDNA libraries by yeast two-hybrid technique using 54 aim protein as a bait. To evaluate the accuracy of the Y2H database, 8 pairs of interactions of the Y2H database were tested by a co-immunoprecipitation assay, and 6 pairs of them were positive. The Y2H database was analyzed by bioinformatics method, estimated the possible co-location place of some proteins, which provided useful information for the next step's further research. On the other hand, we searched 255 pairs of interactions of these 54 proteins in NCBI and HPRD database. We established new mitochondria intermembrane space protein interaction with these 49 pairs of interactions from our previously investigation and combined with 255 pairs of interactions from database and 110 pairs of rotary verification positive interactions. On the basis of the network analysis, we studied the function of SMN1, an interaction protein of COX6B1. The result showed the membrane potential decreased, ATP production increased, ROS level decreased and electron transport chain complex IV activity increased in SMN1 over-expression Hela cells which compared with control cells when treated with 4Gyγradiation. The data showed that SMN1 affected mitochondrion function by interacting with COX6B1 protein.In conclusion, we completed protein interaction network of mitochondria intermembrane proteins, analyzed and verified the data from the network. The primary investigation of SMN1 protein function provides new clues for understanding the function and mechanism of intermembrane space proteins.