1. Why are bacteria called gram-negative or gram-positive? What is special about a gram-negative bacterium’s structure that affects its interaction with an antibiotic? 

Possible student response: Unlike gram-positive bacteria, gram-negative bacteria do not retain a violet color when exposed to a dye called a Gram’s stain. Whereas gram-positive bacteria have only one membrane, gram-negative bacteria have a cell membrane and cell wall. The outer membrane is impermeable to most antibiotics, so even if an antibiotic has the ability to kill gram-negative bacteria, it might not be able to get through the outer membrane to do so.

2. What has been the traditional way to learn how antibiotics cross the bacterial barrier? What situation has provided extra motivation for advances in drug development? 

Possible student response: Traditionally, scientists have studied the bacterial barriers to learn how compounds might cross them. Breaking the tradition, the research summarized in this article focused on the properties of molecules that could help them get across the barrier. Some of today’s research is motivated by the fact that bacteria are becoming resistant to antibiotics, and all of the most critical bacteria on a list created by the World Health Organization are gram-negative bacteria.

3. What is a porin, and why are they important for antibiotic design? 

Possible student response: A porin is a protein that dots the outer membrane of a gram-negative bacterium and acts like a channel, or pore, to allow the cell to take in nutrients. Passage through a porin is typically how an antibiotic can enter gram-negative bacteria.

4. What did chemical biologist Paul Hergenrother and his group do to learn about porin passage?

Possible student response: Hergenrother’s group synthesized 100 compounds that have molecular characteristics similar to those of naturally occurring antimicrobials and then incubated each with
E. coli bacteria in a tube for 10 minutes. Researchers then measured how much of each compound got inside the cells. Next, they analyzed the molecules that had the most success in penetrating the cells, discovering that an amine group was a shared feature.’

5. Once Hergenrother’s group was finished with its initial study of the 100 compounds, what additional studies did the team perform? 

Possible student response: After identifying that an amine group might aid in a molecule’s passage across a bacterial barrier, Hergenrother’s group used a larger set of amine-containing compounds and performed the E. coli incubation test. Using a computer program, the researchers also learned that a rigid, flat structure was more likely to pass through porins than a flexible or spherical one.

6. What antimicrobial did the researchers use to test the new “rules” discovered by their previous experiments? Why did they choose it initially, how was it altered and what was the outcome? 

Possible student response: Researchers used an antimicrobial called deoxynybomycin that is effective only against gram-positive bacteria. Deoxynybomycin is flat and rigid, so it already has “the right geometrical parameters,” said Hergenrother. The researchers synthesized a deoxynybomycin derivative containing an amine group and tested it on gram-negative pathogens. It was effective on all but one pathogen tested.

7. Explain what microbiologist Kim Lewis means when he says this research could “revive the failed effort to rationally design antibiotics.” What fields of science are needed to create designer antibiotics? 

Possible student response: This research shows that existing gram-positive antibiotics may be chemically altered to be effective on gram-negative bacteria. Many biological fields such as microbiology and immunology, as well as chemical fields, such as synthetic organic chemistry and instrumental analyses, are involved in building designer antibiotics.