Translational elongation requires precise coordination between factors involved in releasing one amino acid and adding another to the growing polypeptide chain. This process is coordinated by elongation factors, which support ribosomes in catalyzing the transfer of the peptidyl-tRNA from the P-site to the A-site, where the aminoacyl-tRNA then binds. One such elongation factor is EF-Tu, which is responsible for delivering aminoacyl-tRNAs to the ribosomal A-site. Inhibition of EF-Tu thus leads to inhibition of protein synthesis. The antibiotic Kirromycin is known to specifically target EF-Tu and thereby inhibit translation. Studies reveal that this antibiotic binds to an alpha-helix of EF-Tu, preventing conformational changes that are essential for translocation of the ribosome along mRNA. In Kirromycin-mediated inhibition, the ribosome is unable to bind aminoacyl-tRNA, which thus halts the elongation process of protein synthesis [3]. Kirromycin has proved highly useful as a research tool to study the intricacies of translation and elongation, as well as for development of potential therapeutics aimed at interfering with bacterial protein synthesis [4].

The use of Fusidic Acid

Fusidic acid, another antibiotic, targets another elongation factor, EF-G, which translocates the ribosome along the mRNA transcript. Inhibition of EF-G results in ribosomes stalling at various points along the RNA, thereby affecting the translation of the protein [5]. More recent studies of fusidic acid have revealed that mutations in the bacterial gene that encodes for EF-G leads to resistance to this antibiotic, thus limiting its usefulness in clinical settings [6].

Conclusion

In conclusion, small molecule inhibitors and antibiotics such as Puromycin, Kirromycin, and fusidic acid have been invaluable tools in deciphering the nature of the elongation process in protein synthesis. These inhibitors have provided a means to not only reveal the complexities of translation, but also how its regulation can be disrupted in a variety of diseased states. Through their use, we have gained a greater understanding of the mechanisms that govern protein synthesis and the means to potentially develop new therapeutic interventions towards the treatment of a broad range of disorders.

The progression of the ribosome along the mRNA transcript involves the process of translocation, which is crucial in translational elongation. This process requires GTP hydrolysis, and the involvement of EF-G, a 5 domain G protein and another elongation factor actor. Although EF-TU and EF-G compete for the same binding site on ribosomes, it is the activity of EF-G alone that is responsible for inducing conformational changes in ribosomes to propel them forward. When an aminoacyl-tRNA is delivered into the A site, and EF-TU is released, the ribosome moves three nucleotides along the mRNA transcript, via GTP hydrolysis. This hydrolysis reaction triggers a conformational change in the EF-G protein, which then triggers a shift in the ribosome. The ribosomal subunits move successively via the formation of an intermediate hybrid state, and the importance of the release of EF-G after this was highlighted by the use of fusidic acid, a steroid antibiotic. This chemical compound traps the elongation factor on the ribosome with GDP, thus disabling EF-TU from binding, and preventing another aminoacyl-tRNA from being delivered into the A site of the ribosome. Consequently, translation suffers early termination.

Antibiotics have played a vital role in understanding translational elongation. They include puromycin, which halts the extension of the polypeptide chain, kirromycin, which obstructs the peptide transfer reaction, and fusidic acid, which restrains translocation by freezing the ribosome in position. Through these chemicals' independent inhibitory effects, molecular biologists have expanded their knowledge of the connection between translational dysregulation and neurological function. Comparative translations between healthy and cancerous cells are now better understood. The newly gained knowledge can also help answer more challenging questions, such as the mechanism behind antibiotic resistance in bacteria.

While research is currently ongoing into the structural and functional basis for resistance to kirromycin in mutant EF-TU species of Escherichia coli, there is still much about the mechanism that remains unknown.

Reference:

1. Aviner R. (2020). The science of puromycin: From studies of ribosome function to applications in biotechnology. Comput Struct Biotechnol J. 18:1074-1083. doi:10.1016/j.csbj.2020.04.014.

2. Maracci C et al. (2015). Activities of the peptidyl transferase center of ribosomes lacking protein L27. RNA. 21(12):2047-2052. doi:10.1261/rna.053330.115.

3. Wolf H et al. (1974). Kirromycin, an inhibitor of protein biosynthesis that acts on elongation factor Tu. Proc Natl Acad Sci U S A. 71(12):4910-4914. doi:10.1073/pnas.71.12.4910.

4. Mesters JR et al. (1994). The structural and functional basis for the kirromycin resistance of mutant EF-Tu species in Escherichia coli. EMBO J. 13(20):4877-4885. doi:10.1002/j.1460-2075.1994.tb06815.x.