9. (10 pts) Design an experiment that would test the effect of the presence of t
ID: 276376 • Letter: 9
Question
9. (10 pts) Design an experiment that would test the effect of the presence of the natural substrate, lactose, on the reaction. What would your data look like (you can compare it to data you collected). Include your hypothesis, controls and what different conditions/ratios you are going to test in your experiment (lactose:ONPG) What is your hypothesis? Do you think the absorbance will increase or decrease relative to control in the presence of lactose? Keep in mind why we are not using lactose in the current experiment. a. b. What different ratios of ONPG: lactose are you going to use? c. How your results compare from one ratio to the next? d. What are your controls?Explanation / Answer
Ans:
Experiment: In the o-nitrophenyl ß-galactoside (ONPG) enzyme assay (In presence of fixed conc. of Enzyme and ONPG) add different concentration of Lactose and check the enzymatic activity of ß-galactoside by absorbance.
9a.
Hypothesis: Natural substrate ‘Lactose’ has an effect in the o-nitrophenyl ß-galactoside (ONPG) enzyme assay.
The absorbance value will decrease after addition of lactose in o-nitrophenyl ß-galactoside (ONPG) enzyme assay because lactose act as competitive inhibitor in this assay. Lactose and ONPG both are substrate of ß-galactosidase thus they compete for the active site of the enzyme. Thus amount of ONPG breakdown decrease in comparison to control (where no lactose is used) in presence of lactose results less amount of yellow o-nitrophenol formation provides less absorption value.
9b.
Use lactose maximum concentration which is equal to the ONPG conc. Means 1:1 lactose:ONPG. Other conc. used to 1/10 of the maximum lactose conc. Means. If X?M is the maximum conc. of lactose used then other lactose conc. need to use are X/10?M ,X/100?M, X/1000?M, X/10000?m.
9c. with the decreasing conc. of lactose the absorbance will be increased because the competition is decreased with decreasing conc. of lactose.
9d. Control experiment contains ONPG and ß-galactosidase enzyme. No lactose is required. Thus this set provides the enzymatic activity of uninhibited enzyme.
Related Questions
Navigate
Integrity-first tutoring: explanations and feedback only — we do not complete graded work. Learn more.