13 Jun Streptococcus pneumoniaeis highly dependent upon external sugars
Figure 1.This is the phosphotransferase system that S. pneumoniaeuses for the transport of glucose. (PEP is Phosphoenolpyruvate. HPR is a protein necessary for glucose transport; it also plays a key role in controlling expression of some genes as explained on next page.)
a. What term best describes this transport mechanism? Explain the reason for your answer.
b. What is the immediate source of energy that drives this transport system and allows S. pneumoniaeto scavenge low levels of glucose from its environment?
c. When S. pneumoniaeis growing in the presence of oxygen using glucose as its energy source, which
metabolic pathway (Aerobic respiration, fermentation or anaerobic respiration) would it use? Justify your answer.
d.What is the NET GAIN in ATP generated by S. pneumoniaecatabolizing one molecule of glucose during [Remember to take into account the cost of glucose transport]. Show how you generated this answer.
7.Bill Gates is interested in developing vaccines. His foundation supports the Global Alliance for Vaccine Development (GAVI). He would be very interested in funding projects that provide novel delivery mechanisms for vaccines. Frederica has always wanted to find a way to make a vaccine for Streptococcus pneumoniaethat targets all of the serotypes! This may be her opportunity! Her goal is one vaccine that would be completely protective against all strains of S. pneumoniae. She has an idea to make an edible vaccine that would contain the antigen derived from S. pneumoniae. Frederica decides to try using yogurt as the delivery mechanism! She wants to clone a gene from S. pneumoniaethat encodes an antigen and to express this gene in a microorganism that would grow in yogurt. Eating the yogurt would serve as the vaccine delivery. Consider the various molecules expressed by S.pneumoniae.
a. Choose one molecule that would be an appropriate choice to serve as the immunogen (antigen) for the vaccine preparation. What molecule have you chosen?
b. Explain how your choice would allow an immunized person protection from allstrains of S. pneumoniae.
For the vaccine she must incorporate the gene for the S. pneumoniaeantigen into a bacterium that will survive in the yogurt and will express the gene producing the antigen molecule. One organism found in yogurt is Streptococcus thermophilus. Frederica begins her plan to transform Streptococcus thermophiluswith a plasmid vector encoding the antigen gene.
First Federica must choose a plasmid cloning vector. Frederica understands that a gene encoding resistance to antibiotics is commonly used to select for recombinant organisms (she does not want to include such a gene in her final construct for fear that it would be transferred to normal flora in the gut of the immunized children). As such, her final recombinant S. thermophilusstrain will need some further genetic manipulations. However, for a phase one study, where the vaccine will be tested in the laboratory in animals, it is ok.
c. Prepare a drawing of the cloning vector that you would use. Be sure that your drawing contains all of the essential information needed to demonstrate:
1. That the plasmid will replicate in S. thermophilus
2. Any other genes that you will need to identify your transformants.
d. Prepare a diagram that shows how to construct a recombinant plasmid that would contain the gene that encodes your immunogen.
– Consider the plasmid that you have designed. How will you identify S. thermophiluscells that have been transformed with this plasmid?
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