The Influence On Thylakoid Structure And Photosynthetie Physiology Under Drought Stress And Identification Of Thylakoid Membrane Protein In Nostoc Flagelliforme

Drought stress, one of the major abiotic stress factors, can make an important influence on the growth of plants. As a kind of terrestrial nitrogen-fixing cyanobacterium, Nostoc flagelliforme mainly grow in arid and semi-arid desert steppes. The photosynthesis of N. flagelliforme is vital limited by drought stress. In this study, we analyzed the effects on thylakoid ultrastructure and photosynthetic physiology of N. flagelliforme under the drought stress. In addition, the thylakoid membrane proteins were separated by SDS-PAGE, thylakoid membrane proteins were also identificated by mass spectrometry. The researches have provided the basis for the further study photosynthetic mechanism of N. flagelliforme by drought stress.1. The thylakoid structure and photosynthetic physiology of N. flagelliforme demonstrated significant changes under drought stress. When the moisture content was1194.9%, the thylakoid structure appeared clear. Most of the thylakoid membranes arranged parallelly with cell walls. With the aggravation of drought stress, the structure of thylakoid lamella distorted and gradually changed blurrily and the arrangement tended to be disorder.With the reduction of moisture content in colonies, the contents of Chlorophyll a, allophycocyanin, phycoerythrobilin and phycobilin decreased significantly. The content of carotenoid increased firstly and then decreased, the difference between each other reached an extremely significant level. As the drought stress aggravates, the content of ATP tended to increased and then to decreased. It reached the peak when the water content was51.16%, while it had a rapid decrease when the moisture content reduced from51.16%to7.4%. But ATP content showed no significant difference when the moisture content was1194.9%and7.4%, respectively. Under the drought stress, the content of soluble protein tended to decrease and the difference reached an extremely significant level. The activity of RuBisco firstly increased and it reached the peak when the moisture content was459.2%, and then declined grandually as the drought stress aggravation.2. There were no significant differences between the structure of thylakoids of N. flagelliform which grew under natural environment and the structure of thylakoids of cononies storaged at dry environment for ten years. They both ranged disorderly and the structures were unclear. For ten years dry storaged colonies, there were no net photosynthetic rate, the dark respiration rate declined1400%compare to natural environment colonies. The content of carotenoid remarkablly increased96.58%, while the content of Chlorophyll a, algocyan, allophycocyanin, phycoerythrobilin and phycobilin of cononies storaged at dry environment for ten years decreased at6.17%,248.46%,109.94%,262.19%and56.33%, respectively, compared with the natural environment colonies. But The content of ATP decreased59.4%, and the activities of RuBisco decreased60%in cononies storaged at dry environment for ten years.3. The grinding method and ultrasonic method were applied to break the cells, one-time centrifugation and many times centrifugation step by step to separaet thylakoid crude products, and then the thylakoid membranes were purified in gradient density sucrose by high speed centrifugation, the thylakoid membrane proteins were extracted and separated by SDS-PAGE. The thylakoid membrane proteins were identified by Matrix-assisted laser desorption ionization-time of flight-tandem mass spectrometry （MALDI-TOF-TOF/MS）. The results showed that14bands were separated in the image of thylakoid membrane protein of natural N. flagelliform by DS-PAGE electrophoresis, their mass weight are mainly from25kD to60kD. Of the14protein bands,8protein bands were identified by database search. Based on their biological functions, the thylakoid membrane proteins were categorized into4groups:①Unknown protein:hypothetical protein Npun_R1321, hypothetical protein Npun_R3785, Hypothetical protein N9414₀2186;②Metabolic enzymes Photosystem II manganese-stabilizing protein PsbO, ATP synthase subunit alpha:F1F0ATP synthase subunit alpha, FiF0ATP synthase subunit beta;③Domain protein:hemerythrin HHE cation binding domain-containing protein;④Carbohydrate-selective porin OprB. They might play an important role in the photosynthesis of N. flagelliforme.