Signal Response Of Yeast Cell To Pb And Cr Stress And Biosorption Polluted Waste Water By Yeast

With the rapid growth of industry,the content of Pb and Cr???rapidly increases in the environment.Acturally,Pb and Cr???is harmful for the human being,animals,plants and microorganisms.The toxicant mechanism of Pb and Cr???has caused wide public concern all over the world.Recently studies on the toxicant mechanism of Pb and Cr???mainly focuse on the animals and plants,but little attention and study paid on microorganisms.Microorganisms plays an important role in recycling nutrients,plant growth and development and controlling environment pollution,and so on.Therefore,to study the toxicant effect of Pb and Cr???on the microorganisms,thus proposing concrete measures for enhancing the tolerance of microorganisms against heavy metals andimprovingtheabilityofmicroorganismsremovingheavymetals.The research result of this field has important practical significance and theoretical value to the environmental protection.Therefore,this present paper regards yeast cells as the research object.In present study,we make research on the toxicant effect and mechanism of Pb and Cr???on the microorganisms,the effect of metallothionein expression in yeast on the toxicity of Pb and Cr???,to compare the removal effects of transgenic MT yeast and host strain on Pb and Cr???.Main results for this study are as follows:1.The results reveal that the cellular bioactivity of yeasts is inhibited by lead nitrate at the concentration from 0 to 100 mg/L and cell death induced by Pb is found.Yeast cell mortality increases with the increase of Pb concentration and the extension of time.Cell death induced by Pb is decreased by Z-Asp-CH2-DCB.Meanwhile,the levels of ROS and Ca²⁺elevate significantly and the mitochondrial membrane potential decrease dramatically in yeast cells under Pb stress.The content of intracellular cyt c elevate significantly in yeast cells under Pb stress.Cell death induced by Pb is decreased by ascorbic acid?AsA?at the concentration of 1 mM and is inhibited by the calcium chelator?EGTA?at the concentration of 0.5 mM or the specific inhibitor of plasma membrane Ca²⁺channel?LaCl₃?at the concentration of 0.1 mM.In addition,the levels of intracellular Ca²⁺induced by Pb is decreased by AsA.The results show that cell death induced by Pb is related to the levels of intracellular ROS and Ca²⁺.The opening of mitochondrial membrane permeability transition pore channels may be induced by high levels of Ca²⁺,or mitochondrial membranes are broken by high levels of ROS,which lead to decreasing membrane potential and cause cell death by activating down-stream relevant signal pathways.Nitric oxide?NO?have many regulatory roles in response to abiotic stresses and play important roles in the regulation of cell death.In order to study the role of nitric oxide on cell death induced Pb,we test the effect of Pb on NO level in yeast.The result show that the levels of NO elevated significantly in yeast cells under Pb stress.Cell death induced by Pb is decreased by nitric oxide scavenger cPTIO or nitric oxide synthase inhibitor L-name.Cellular toxicity induced by Pb is increased by nitric oxide donor SNP.To further investigate the mechanisms of nitric oxide regulation in cell death induced by Pb,we test the effect of nitric oxide on Pb content in yeast.The result show that the intracellular Pb content is inhibitor by cPTIO or L-name.Instead,the intracellular Pb content is enhanced by SNP.Based on these findings,we assume nitric oxide may promote intracellular Pb transport in Pb-induced yeast cell death.In coclusion,on the one hand,cell death induced by Pb is related to the levels of intracellular ROS and Ca²⁺.The opening of mitochondrial membrane permeability transition pore channels may be induced by high levels of Ca²⁺,or mitochondrial membranes are broken by high levels of ROS,which lead to decreasing membrane potential and cause cell death by activating down-stream relevant signal pathways.On the other hand,nitric oxide may promote intracellular Pb transport in Pb-induced yeast cell death.2.The results reveal that the cellular bioactivity of yeasts is inhibited by Cr???at the concentration from 0 to 7 mM and cell death induced by Cr???is found.The levels of ROS,Ca²⁺and NO elevated significantly and the mitochondrial membrane potential decreased dramatically in yeast cells under Cr???stress.Cell death induced by Cr???is decreased by ascorbic acid?AsA?at the concentration of 1 mM or catalase?CAT?at the concentration of 200 U.Cell death induced by Cr???is decreased by the calcium chelator?EGTA?at the concentration of 0.5 mM or the specific inhibitor of plasma membrane Ca²⁺channel?LaCl₃?at the concentration of 0.1 mM.Cell death induced by Cr???is decreased by nitric oxide scavenger cPTIO or nitric oxide synthase inhibitor L-name.In addition,the levels of intracellular Ca²⁺induced by Cr???is decreased by cPTIO,instead,the levels of intracellular NO induced by Cr???can not be decreased by EGTA.The results show that cell death induced by Pb is related to the levels of intracellular ROS,NO and Ca²⁺.The opening of mitochondrial membrane permeability transition pore channels may be induced by high levels of Ca²⁺,which lead to decreasing membrane potential and cause cell death by activating down-stream relevant signal pathways.3.On the basis of transgenic MT yeast,we test the effect of MT on cell death induced Pb and Cr???.The results show that the transgenic MT yeast death rate is significantly decreased compared to the host strain.The levels of ROS and Ca²⁺of the transgenic MT yeast is significantly decreased compared to the host strain.The results show that cell death induced by Cr???and Pb is decreased by MT,due to the levels of intracellular ROS and Ca²⁺is decreased by MT.4.Biosorbent surface morphology and material structures is studied using SEM and FTIR.The outline of the yeast is oblong shape,with smooth surface and high superficial area,which was advantageous to remove Cr???.EDX analysis has provided the direct evidence for the specific adsorption of Cr???ions onto the surface of the host and M1strains.FTIR result show that hydroxyl,amine,phosphoric acid and amide groups all play a role in Cr???adsorption.The effect of the dose,initial pH value,temperature,time,Cr???initial concentration is researched at the index of Cr???removal rate.The amount of Cr???adsorbed by the yeast increase sharply at the beginning,and then gradually reach equilibrium after 30 min.The results show that the best removal condition is that the initial pH value of 1.The amount of Cr???adsorbed by the yeast decrease with pH increase.The amount of Cr???adsorbed by the yeast is increased with temperature from 20 to 40?.The amount of Cr???adsorbed by the yeast is decreased with dose increase.The maximum biosorption is observed at 8.01 mg/g using a 0.5 g/L dose.The biosorption capacity of Cr???by the M1strain increases with an increase in the initial Cr???concentration.The result show that the Langmuir and Freundlich isotherm model can better optimize the experimental data.The saturated monolayer sorption capacity increases from 2.6 to 8.2mg/g at 30?after MT expression.The square of the correlation coefficient?R²=0.9691?of the pseudo-second-order equation is better than the value for the pseudo-first-order equation.The interaction between pH and dosage is significant.The interaction between pH and temperture is not significant.The interaction between temperature and dosage is not significant.The optimal conditions for removal Cr???are pH 3.7,a biosorbent dosage of2.02 g/L,and temperture 29.5?.The maximum predicted value of removal Cr???is 6.49mg/g.Under optimal conditions,the genetic engineerd strain is applied to treatment electroplating chromium-containg wastewater.The result show that the content of Cr???in the electroplating wastewater is below 0.5 mg/L after 3-batch treatment with M1 strain.5.The effect of the dose,initial pH value,temperature,time,initial Pb concentration is researched at the index of Pb removal rate.The amount of Pb adsorbed by the yeast increased sharply at the beginning,and then gradually reach equilibrium after 20 min.The amount of Pb adsorbed by the yeast is decreased with pH increase.The maximum biosorption is observed at pH=5.The amount of Pb adsorbed is increased from 5.6 to 48.3mg/g with an increase in initial Pb concentration from 10 to 90 mg/L.The amount of Pb adsorbed by the yeast is decreased with dose increase.The amount of Pb adsorbed is increased from 30.6 to 37.2 mg/g with temperature from 20 to 40?.The result show that the Langmuir and Freundlich isotherm model can better fit the experimental data.The saturated monolayer sorption capacity increases from 125 to 312.5mg/g at 30?after MT expression.The square of the correlation coefficient?R²=0.9491?of the pseudo-second-order equation is better than the value for the pseudo-first-order equation.