| Mitochondria, which provides energy for the cells of the body, is the main place to cellular ATP synthesis and oxidative phosphorylation. Besides, mitochondria play an important role in apoptosis and cellular signal transduction, etc. Mitochondria structures with double membrane. The natural mitochondrial transmembrane potential is prerequisite for the normal physiological function of mitochondria. When ATP synthase supply energy occurs with obstacles, mitochondrial transmembrane potential will decrease and with it a series of changes will happen in cell. Mitochondrial permeability transition pore which is considered to be one of the key factors that lead to cell apoptosis is a special channel in mitochondria. CyPD is considered to be a composition of MPTP and can take part in apoptosis which induced by the open of MPTP. The exact molecular structure of MPTP is unclear.There is still a controversy that whether it only appeares in pathological situation or whether the MPTP plays a natural physiological function in maintaining the stability of mitochondria and cell. Mitchondria can produce NO which has a anti-oxidation effect in low concentration. NO as a bioactive substances can also regulate oxygen consumption and the generation of ROS. Fruit treated with NO can prolong the storage time and keep the fruit quality. While there were few study about the change of mitochondrial permeability of fruit treated with NO and the regulation of NO to CyPD.In this study, CyPD of ‘FeiCheng' peach was studied by gene cloning and prokaryotic expression included PCR technique,RACE technique and protein expression technique system. Yeast two hybrid between ANT and CyPD was tested. Mitochondrial permeability relevant indexes(ATP synthase, cytochrome c, mitochondrial membrane potential, mitochondrial membrane permeability,etc) of ‘Feicheng' peach treated by 5 ?mol L-1, 15 ?mol L-1, 15 ?mol L-1 NO, c-PTIO and control were tested.The experimental results showed that the length CyPD gene was 663 bp, of which 615 bp encoding amino acid. CyPD protein was consist of 204 amino acid. CyPD of persica and malus were closer than other species in analysis of phylogenetic tree. CyPD could take part in reacting with nitrosation and glutathionylation. It indicated that the part of extra membranes of CyPD and ANT didn't react with each other in Yeast two hybrid test. NO had a facilitation in declining the copy number of CyPD gene. This result showed that NO could take part in regulating MPTP by the way of declining the copy number of CyPD gene. The content of cytochrome c of peach treated with NO decline during storage, and it treated with c-PTIO(elimination NO) would increase. The activity of ATP synthase of peach treated with NO could enhance during storage, of which 15 ?mol L-1 NO treatment had the best effect. As the the extension of storage time, mitochondrial permeability of different treatment increase in different degrees, of which the mitochondrial permeability of peach treated with 5 ?mol L-1 NO maintained at low level in the whole storage. NO could enhance the capacity of resistance of mitochondrial membrane to stress signals, while elimination of NO would have the opposite effect. After adding ruthenium red, the fluorescence intensity of all different treatment mitochondria declined, of which 15 ?mol L-1 NO treatment had the largest decline in value. This experiment showed that NO had a role in regulating calcium regulatory protein of mitochondria. |
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