Columbia College BIOL BIOL W3041 Questions and Answers > Chloroplasts contain six compartments - outer membrane, intermembrane space.

Document Content and Description Below

Chloroplasts contain six compartments - outer membrane, intermembrane space, inner membrane, stroma, thylakoid membrane, and thylakoid lumen (Figure 1) -each of which is populated by specific sets of ... proteins. To investigate the import of nucleus‐encoded proteins into chloroplasts, you have chosen to study ferredoxin (FD), which is located in the stroma, and plastocyanin (PC), which is located in the thylakoid lumen, as well as two hybrid genes: ferredoxin with plastocyanin signal peptide (pcFD) and plastocyanin with ferredoxin signalpeptide (fdPC). You translate mRNAs from these four genes in vitro, mix the translation products with isolated chloroplasts for a few minutes, re‐isolate the chloroplasts after protease treatment, and fractionate them to find which compartments the proteins have entered (Figure 2A). The status of the normal and hybrid proteins at each stage of the experiment is shown in Figure 2B: each lane in the gel corresponds to a stage of the experiment as indicated alongside the experimental protocol in Figure 2A. a. How efficient is chloroplast uptake of ferredoxin and plastocyanin in your in vitro system? How can you tell? b. Are ferredoxin and plastocyanin localized to their appropriate chloroplast compartments in these experiments? How can you tell? c. Are the hybrid proteins imported as you would expect if the N‐terminal signal peptides determined their final location? Comment on any significant differences. d. Why are there three bands in experiments with plastocyanin and pcFD but only two bands in experiments with ferredoxin and fdPC? To the extent you can, identify the molecular species in the bands and their relationship to one another. e. Based on your experiments, propose a model for the import of proteins into the stroma and thylakoid lumen. Figure 1. The six compartments of a chloroplast. Expand image Figure 2. Import of ferredoxin and plastocyanin into chloroplast compartments. (A)Experimental protocol. (B) Gel analysis. Samples from each stage in the experimental protocol were analyzed by gel electrophoresis. Each lane in (B) corresponds to a particular experimental treatment in (A). Ferredoxin gene segments are black; plastocyanin genesegments are white. The signal peptides in all genes are located at the left (N‐terminal) end. The ferredoxin signal peptide in the hybrid gene is shown as fd; the plastocyanin signal peptide in the hybrid is shown as pc. Please explain the answers fully. I want to be able to understand it because the answer key does not make complete sense to me. Thank you:) [Show More]

Last updated: 1 year ago

Preview 1 out of 3 pages