Teixobactin Synthesis Lab Report

In the 1940s antibiotics were widely introduced as a cure for common types of diseases.
However, some of the diseases started to build up a resistance to the antibiotics and in result, the antibiotics became ineffective. In order to help resolve the crisis, antibiotics were found through screenings of soil microorganisms. However, soil microorganism’s antibiotics were depleted by the 1960s and their antibiotic effects were unable to be replicated through synthesis. In this experiment, the researchers developed numerous methods to cultivate uncultured organisms in their environment. The goal of this experiment was to find antimicrobials in the uncultured soil. Through this successful experiment, the researchers were able to discover a new antibiotic
that showed no detectable signs of resistance. The cultivation technique used in the experiment relied on a multichannel device called the iChip. To perform this technique, it was crucial that each channel in the iChip contained only one bacterial cell. In order to do this, a soil sample was diluted, housed between two semi-permeable membranes, and then placed back into the soil that it originated from. This created a diffusion process that pumped growth factors and nutrients through the chambers into the isolated soil sample allowing the isolated uncultured bacteria to grow. After a colony was produced it was screened for antimicrobial activity on plates containing S. aureus. The screening research found good activity from a beta-proteobacteria named Eleftheria terrae, so the researchers ran a genome scan on the organism followed by PCR. However, 16S rDNA and in silico DNA/DNA hybridization showed the researchers that the organism was related to the known genus Aquabacteria, a Gram-negative organism, which is not an antibiotic producer. Conversely, when the researchers used mass spectrometry on the sample, they found that part of it had an unusual molecular mass of 1,242 Da. This portion of the sample was isolated and the structure was looked at closer using NMR spectroscopy and advanced Marfey’s analysis. Through these techniques they were able to find out that the isolated part of the molecule is a depsipeptide composed of enduracididine, methylphenylalanine, and four D-amino acids. A homology search also identified that the molecule contained two non-ribosomal large peptide synthetase coding genes called txo1 and txo2. The researchers that discovered this new antibiotic named it teixobactin.
After further research, it was determined that teixobactin had great activity against Gram-positive pathogens and even strains of bacteria that were drug-resistant.
Teixobactin was also very active against Clostridium difficile and Bacillus anthracis, while also showing stronger bactericidal activity against S. aureus than the antibiotic vancomycin did. This indicated that teixobactin is a very strong antibiotic. On the other hand, most Gram-negative bacteria were not affected by teixobactin with the exception of E. coli asmB1. When further research was done, it was discovered that teixobactin is not harmful to mammalian NIH/3T3 and HepG2 cells, even when tested at its highest dose. Therefore, teixobactin could be used to treat mammalian diseases. Evidence that supported teixobactin was a peptidoglycan synthesis inhibitor was found when the researchers ran tests using label incorporation in the bacteria S. aureus. The tests confirmed that teixobactin inhibited synthesis of peptidoglycan, however it had no effect on DNA, RNA, or protein. Due to resistance not being a factor, this suggested to the researchers that the teixobactin may bind to a number of targets, but it will not bind to a protein. The researchers then discovered that teixobactin was able to bind to modified forms of lipid II and to undecaprenyl-pyrophosphate. This showed that teixobactin reacts with the peptidoglycan precursor and not with one of the enzymes. Another important piece of information that the
researchers presented was that teixobactin has great lysis ability when bound to a WTA precursor; this is primarily due to its lack of a cell wall due to digestion. Ultimately, this successful experiment tested to see if uncultured bacteria contained any new sources of antimicrobials and what was found was teixobactin, a promising therapeutic candidate.