BIOL-M 350 Lecture Notes - Lecture 3: National Center For Biotechnology Information, Lactobacillus Acidophilus, Human Microbiota
M350 Homework 3
Questions based on: McKinney, JD, et al. 2000. Persistence of Mycobacterium tuberculosis in macrophages
and mice requires the glyoxylate shunt enzyme isocitrate lyase. Nature. 406: 735-738.
For questions answered with sentences, use your own words. You may use additional resources (e.g.,
textbooks, primary literature and reviews, and websites such as Metacyc, BRENDA enzymes, KEGG) to help
you answer these questions. Demonstrate your understanding using clear and concise language. Points
may be deducted for answers that are phrased in a vague or confusing manner.
1. Fatty acids are hypothesized to be an important carbon/energy source during M. tuberculosis lung
tissue infections. Fatty acids are degraded via beta-oxidation and the glyoxylate shunt. ***Consider
how many carbons are in a fatty acid (e.g., palmitate) and explain why the glyoxylate shunt is
required to grow on fatty acids in two sentences or less.
Fatty acids have >4 carbons, but their degradation by beta-oxidation results into units of acetyl-CoA.
Acetyl-CoA must be processed by the glyoxylate shunt to keep the two carbons in the acetyl-group of
acetyl-CoA for biosynthesis, lest they be lost as CO2 in the lower TCA cycle.
2. In a two sentences or less, what is the hypothesis being tested in Fig 2a and how was it tested?
The hypothesis is that the glyoxylate shunt is important for survival in lung tissues. To test the hypothesis
they compared survival of the wild-type versus ICL mutant strains in infected mouse lungs over time.
3. Define these terms as they relate to bacterial genetics (stick to what we’ve covered in class; beware
of online definitions referring to things like combinations of homozygous recessive mutations):
a. Polar effects – mutation in an upstream stream disrupts the normal expression of
downstream genes – usually occurs when genes are in an operon and thus transcribed on a
common transcript
b. Complementation – a wild-type phenotype is restored when the wild-type gene is expressed
from an ectopic location (usually from a plasmid). WARNING –definition online often apply
specifically to diploids. Most bacteria are not polyploidy and therefore these explanations do
not apply. Combining two gene deletions /mutations cannot restore a wild-type gene.
c. Empty vector control - the same vector/plasmid used to complement the mutation is
introduced but does not contain the wild-type gene. This ensures that the restoration of the
phenotype is due to the reintroduction of the wild-type gene and not some other sequence on
the plasmid.
4. In three sentences or less, STATE and EXPLAIN the hypothesis being tested in Fig 2b. In your
answer, use the above terms from #3 (or related terms).
The hypothesis was that the lower mutant survival in Fig 2a was due to the loss of the ICL gene and not due
to polar effects on nearby genes. To test this hypothesis they compared survivability of the mutant
expressing the ICL gene from a plasmid versus the mutant with an empty vector control. The empty vector
was important to show that restoration of survivability is due to the expression of ICL and not due to some
other factor on the plasmid.
5. In one sentence, what is the hypothesis being tested in Fig. 2e?
The hypothesis was that isocitrate lyase was required to kill mice in an infection model.
find more resources at oneclass.com
find more resources at oneclass.com
Document Summary
Persistence of mycobacterium tuberculosis in macrophages and mice requires the glyoxylate shunt enzyme isocitrate lyase. For questions answered with sentences, use your own words. You may use additional resources (e. g. , textbooks, primary literature and reviews, and websites such as metacyc, brenda enzymes, kegg) to help you answer these questions. Demonstrate your understanding using clear and concise language. Points may be deducted for answers that are phrased in a vague or confusing manner: fatty acids are hypothesized to be an important carbon/energy source during m. tuberculosis lung tissue infections. Fatty acids are degraded via beta-oxidation and the glyoxylate shunt. ***consider how many carbons are in a fatty acid (e. g. , palmitate) and explain why the glyoxylate shunt is required to grow on fatty acids in two sentences or less. Fatty acids have >4 carbons, but their degradation by beta-oxidation results into units of acetyl-coa.
