Study Of Effects Of Protein S-nitrosylation On The Development Of PSE Pork

Due to pale color,soft texture and exudative appearance,pale,soft and exudative(PSE)pork declines the purchasing desire of consumers.Moreover,the poor processing ability and low yield caused by protein denaturation have increasingly limited the development of meat industry and become the urgent problem.Nowadays,it has been widely recognized that the major cause of PSE pork is high glycolysis rate at early post-mortem stage while the muscle temperature is still high.Most researches have focused on pre-slaughter factors while less post-slaughter factors have been studied.Increased reactive oxygen and nitrogen species would produce during post-mortem aging.Nitric oxide(NO)could modify protein cysteine to form protein S-nitrosothiol regulating protein structure,activity,function and interaction.Protein S-nitrosylation could modulate muscle metabolic function during post-mortem aging and eventually affect pork quality.Thus,it is significant to explore the effects and mechanism of protein S-nitrosyaltion on the development of PSE pork.Our study focused on the effects of S-nitrosylation on calcium transfer proteins and glycolytic enzymes in order to explain the mechanism of PSE pork development from those two metabolic pathways including calcium release and glycolysis.The details and results are as follows:1.The comparison of total protein S-nitrosylation level between PSE and RFN porkThe total protein S-nitrosylation level between PSE and red,firm and non-exudative(RFN)pork was compared in this chapter.Seven PSE and seven RFN pork longissimus thoracis(LT)muscles were chosen according to pH and L*at 1 h and drip loss at 24 h post-mortem.The results showed that pH,L*at 1 h and 24 h,and drip loss at 24 h post-mortem were significantly different between PSE and RFN pork(P<0.05).PSE pork possessed a higher shear force at 24 h post-mortem than RFN pork(P<0.05).At 1 h post-mortem,lower nitric oxide synthase(NOS)activity and higher neuronal nitric oxide synthase(nNOS)expression were detected in PSE pork than in RFN pork(P<0.05).In addition,as shown in SDS-PAGE of total protein S-nitrosylation,S-nitrosylated proteins distributed in different molecular weight especially from 15 to 55 kDa.The band quantities of total nitrosylated proteins had no significant differences between those two groups while some variations of band intensity were observed.These results indicate that total protein S-nitrosylation level was significantly different between PSE and RFN pork and protein S-nitrosylation level varied based on protein species,which underlies the different S-nitrosylation level of specific proteins between PSE and RFN pork.2.Study of effect of protein S-nitrosylation on calcium homeostasisThe aim of this chapter was to explore the effects of protein S-nitrosylation on calcium homeostasis during post-mortem aging.LT muscles at 1 h post-mortem were chosen based on the classfication of first chapter.Sarcoplasmic calcium concentration,sarcoplasmic reticulum calcium ATPase(SERCA)activity,the expression and S-nitrosylation level of SERCA1 and ryanodine receptor 1(RyR1)were determined.Results showed that PSE pork possessed a higher sarcoplasmic calcium concentration than RFN pork(228.33 versus 38.89 ?M;P<0.05).In PSE pork,the expression of RyR1 and SERCA1 was lower than that in RFN meat,while the relative S-nitrosylation level of RyR1 and SERCA1 was higher(P<0.05).In addition,a lower activity of SERCA was detected in PSE pork compared to RFN pork(P<0.05).These results indicate that S-nitrosylation of RyR1 could activate RyR1 to accelerate sarcoplasmic calcium release,and S-nitrosylation of SERCA 1 could inactivate SERCA1 to reduce calcium uptake in sarcoplasm.Thus S-nitrosylation of RyR1 and SERCA1 could exert an important effect on modulating the calcium concentration in sarcoplasm.3.Study of effect of protein S-nitrosylation on glycolytic enzymesThis chapter studied the effect of protein S-nitrosylation on the development of PSE pork by comparing the activity,expression and S-nitrosylation level of key metabolic enzymes between RFN and PSE pork.LT muscles at 1 h postmortem were chosen and glycogen content,lactic acid content and the activity,expression and S-nitrosylation level of glycogen phosphorylase(GP),phosphofructokinase(PFK)and pyruvate kinase(PK)were determined.Results showed that PSE pork had considerably lower glycogen and higher lactic acid content compared with RFN pork(P<0.05).Compared to PSE pork,RFN pork had lower activities and higher S-nitrosylation levels of GP,PFK and PK(P<0.05).Moreover,GP expression in RFN pork was lower(P<0.05)while no significant differences of PFK and PK expression were observed between these two groups(P>0.05).These results indicate that higher S-nitrosylation levels of GP,PFK and PK correspond with lower glycolytic enzymes activities.Abnormally high S-nitrosylation levels of GP,PFK and PK could partly inhibit enzyme activities.Therefore,protein S-nitrosylation could regulate postmortem glycolysis and meat quality by modulating enzyme activities.