They thought they were adding in the correct concentration of co-substrates. How
ID: 144611 • Letter: T
Question
They thought they were adding in the correct concentration of co-substrates. However, instead of adding in 10 mg/mL (14.6 mM) of NAD+ and 10 mg/mL 14.1 mM) NADH, the student added 0.1 mM NAD+ and 0.1 mM NADH from part A of the practical. They completed the assay and measured the absorbance of their systems. The following data obtained and plotted.
i) Describe the trend in the results as depicted in the above graph.
ii) Describe the trend that would have been observed if the correct NAD+ and NADH solutions would have been used and iii) what has gone wrong in the experiment. In your answer describe the reaction catalysed by lacatate dehydrogenase and the change in substrate and products that should have occurred. Include which species is oxidised and reduced, and thus, the form of the co-substrate required for the reaction. (Hint: think about what is being measured in this part of the practical and how the reaction is catalysed by lactate DH)
Part C: Co-substrate requirements of lactate DH 1,01 0.8 p 0.6 0.4 0.2 0.0 NADH NAD+ NoneExplanation / Answer
Hi,
in the above experiment, lactate was catalysed by LD to produce pyruvate. Pyruvate reacts with hydrazine reagent to give a colored compound which is measured at 510nm.
The give graph indicates that absrobance or enzyme activity remains same when NAD+ is added and when nothing is added. Enzyme activity reduces slightly when NADH is added.
The presence of NAD+ drives the reaction forward producing more absorbance. if NADH is added in excess then revese reaction would have been favored producing less of colored substance. If correct amount was added, it would have given rise to higher moles of NAD+ than the NADH. That would have created more absorbance compared to none and NADH suppliments. When NAD+ is added, it would reduce to yield NADH. if NADH is getting catalysed then it would oxidise to give you NAD+.
Related Questions
drjack9650@gmail.com
Navigate
Integrity-first tutoring: explanations and feedback only — we do not complete graded work. Learn more.