Explain and discuss the results shown for each glucose so rate of oxygen uptake and rate of sugar uptake calculations. Document Preview: BS2001 –Metabolism XXXXXXXXXXUptake and metabolism of glucose...

BS2001 –Metabolism 2013-03-05 Uptake and metabolism of glucose and glucose analogues by yeast. In this practical you will compare D-glucose, L-glucose, 2-deoxy-D-glucose and ethanol as substrates for aerobic respiration by yeast. The ability of yeast to oxidise these substrates will be measured using an oxygen electrode. If the yeast are unable to oxidise any of these substrates it may be because:- The yeast is unable to take up the substrate from the medium, or, The yeast cannot metabolise the substrate once it has been taken up. You can distinguish between these alternatives by measuring the rate of uptake of the three sugars. Sugar uptake will be measured by measuring the decrease in sugar concentrations in the medium over a period of time. All three sugars are reducing sugars and can be measured by the DNSA reaction. Method You are provided with a 3g/100 mL suspension of yeast in 20mM phosphate buffer pH 7.5. To measure the rates of oxygen uptake set up the following in the oxygen electrode:- 2.9 mL Kphos buffer ph 7.5 1 mL Yeast suspension mL 50 mM substrate (sugars) Measure the rate of oxygen consumption by writing down the oxygen concentration in your lab book every 15 sec over a 4 min period. You will also need to measure the rate of oxygen consumption using water instead of sugar (why?) Calculate the rate of oxygen uptake a umol/min/g yeast suspension, assume oxygen solubility of 0.25 umol/mL. Sugar uptake. Set up the following reactions: 2.9 mL Kphos buffer pH 7.5 1 mL yeast suspension mL 50 mM sugar Immediately remove 1 .5 mL into a centrifuge tube. Pellet t the yeast and remove 1 mL of supernatant. Incubate the remaining mixture at room temperature for 30 min. Remember to shake the mixture occasionally. After 30 min remove a second 1.5 mL from each reaction, centrifuge the yeast out and retain the 1 mL of supernatant. Add 1 mL of DNSA to each of the supernatants and place into a boiling water bath for 5 min. Cool under a tap and measure the...