Flashcards in MIDTERM 2 - Indole and Catabolite Repression Deck (19):
What is tryptophan metabolism a critical diagnostic for?
Distinguishing between Escherichia from other members of the Enterobacteriaceae
Medium containing proteins digested with trypsin and rich with many amino acids especially tryptophan.
What must bacteria do in order to metabolize tryptophan from the environment?
the bacteria must have a way to translocate the tryptophan into the cell
What happens to tryptophan once it enters the cell?
Its hydrolyzed by tryptophanase.
2. this produces indole + ammonia + pyruvate
3. Indole and Ammonia results from removal of nitrogen in the amino acid
4. Pyruvate being part of the carbon backbone
Out of the three products ammonia, pyruvate, and indole, which one is used for production of energy?
Where is tryptophanase production highest in?
Tryptophanase production and activity is highest in cells growing AEROBICALLY.
True or False: is tryptophan metabolism a fermentative process?
What can inhibit the pathway of tryptophanase
The presence of Glucose
How is the inhibition of the tryptophanase pathway made possible
Through the inhibition of the production of alternative enzymes by an unknown catabolite of glucose metabolism (catabolite repression)
What happens to indole in presence of glucose?
Level of indole production drops below detectable levels even for organisms capable of tryptophan metabolism
How do you get optimal evaluations of the ability to produce indole?
1. Cultures grown aerobically in tryptone
2. No glucose in tryptone broth
3. Incubated at organisms preferred growth temp
4. for 48 hours
What reagent do you add after incubating your organism in tryptone?
What is kovacs reagent?
1. acidified p-DMAB and
2. Amyl Alcohol
Significance of p-DMAB and its flaws?
It reacts with indole to produce a red-violet product called rosindole.
2. Unfortunately p-DMAB reacts with tryptophan as well to produce the same product. This gives rise to erroneous interpretation.
Why is amyl Alcohol important in the Kovacs reagent?
it serves to separate the substrate tryptophan from the product indole
Why is Amyle Alcohol suited for this test?
1. Indole is soluble in amyl alcohol but tryptophan is not
2. p-DMAB is soluble in it
3. Amyl alcohol is immiscible in water and therefore separates from it
4. Amyl alcohol is less dense than water and forms an easily visible layer on top of the aqueous culture medium
Positive Reaction for Indole test?
Dark Red-Violet color in the amyl-alcohol layer on top of the broth
Negative reaction for indole test?
Any color other than dark Red-violet (pink, Orange, Or Amber) in the amyl-alcohol layer on top of the broth.