Puromycin is an

antibiotic An antibiotic is a type of antimicrobial substance active against bacteria. It is the most important type of antibacterial agent for fighting pathogenic bacteria, bacterial infections, and antibiotic medications are widely used in the therapy ...

protein synthesis inhibitor which causes premature chain termination during

translation Translation is the communication of the semantics, meaning of a #Source and target languages, source-language text by means of an Dynamic and formal equivalence, equivalent #Source and target languages, target-language text. The English la ...

Inhibition of translation

Puromycin is an aminonucleoside

, derived from the '' Streptomyces alboniger'' bacterium, that causes premature chain termination during translation taking place in the ribosome. Part of the molecule resembles the 3' end of the aminoacylated tRNA. It enters the A site and transfers to the growing chain, causing the formation of a puromycylated nascent chain and premature chain release. The exact mechanism of action is unknown at this time but the 3' position contains an amide linkage instead of the normal

ester In chemistry, an ester is a compound derived from an acid (either organic or inorganic) in which the hydrogen atom (H) of at least one acidic hydroxyl group () of that acid is replaced by an organyl group (R). These compounds contain a distin ...

linkage of tRNA. That makes the molecule much more resistant to

hydrolysis Hydrolysis (; ) is any chemical reaction in which a molecule of water breaks one or more chemical bonds. The term is used broadly for substitution reaction, substitution, elimination reaction, elimination, and solvation reactions in which water ...

and stops the ribosome. Puromycin is selective for either

prokaryotes A prokaryote (; less commonly spelled procaryote) is a single-celled organism whose cell lacks a nucleus and other membrane-bound organelles. The word ''prokaryote'' comes from the Ancient Greek (), meaning 'before', and (), meaning 'nut' ...

or eukaryotes. Also of note, puromycin is critical in mRNA display. In this reaction, a puromycin molecule is chemically attached to the end of an

mRNA In molecular biology, messenger ribonucleic acid (mRNA) is a single-stranded molecule of RNA that corresponds to the genetic sequence of a gene, and is read by a ribosome in the process of Protein biosynthesis, synthesizing a protein. mRNA is ...

template, which is then translated into protein. The puromycin can then form a covalent link to the growing peptide chain allowing the mRNA to be physically linked to its translational product. Antibodies that recognize puromycylated nascent chains can also be used to purify newly synthesized polypeptides and to visualize the distribution of actively translating ribosomes by immunofluorescence.

Peptidase Inhibitor

Puromycin is a reversible inhibitor of dipeptidyl-peptidase II ( serine peptidase) and cytosol alanyl aminopeptidase ( metallopeptidase). The mechanism of inhibition is not well understood, however puromycin can be used to distinguish between aminopeptidase M (active) and cytosol alanyl aminopeptidase (inhibited by puromycin).

Cell culture

Puromycin is used in cell biology as a selective agent in cell culture systems. It is toxic to prokaryotic and eukaryotic cells. Resistance to puromycin is conferred by the ''pac'' gene encoding a puromycin ''N''-acetyl-transferase (PAC) that was found in a Streptomyces producer strain. Puromycin is soluble in water (50 mg/mL) as colorless solution at 10 mg/mL. Puromycin is stable for one year as solution when stored at -20 °C. The recommended dose as a selection agent in cell cultures is within a range of 1-10 μg/mL, although it can be toxic to eukaryotic cells at concentrations as low as 1 μg/mL. Puromycin acts quickly and can kill more than 99% of nonresistant cells within one day.

Selection of ''Escherichia coli''

Puromycin is poorly active on ''E. coli''. Puromycin-resistant transformants are selected in LB agar medium supplemented with 125 μg/mL of puromycin. But use of puromycin for ''E. coli'' selection requires precise pH adjustment and also depends on which strain is selected. For hassle–free selection and optimum results the use of specially modified puromycin is possible. Plates containing puromycin are stable for 1 month when stored at 4 °C.

Selection of yeast

Puromycin resistance in yeast can also be conferred through expression of the puromycin N-acetyl-transferase (''pac'') gene. Lethal concentrations of puromycin are much higher for strains of ''Saccharomyces cerevisiae'' than mammalian cell lines. Deletion of the gene encoding the multidrug efflux pump Pdr5 sensitizes cells to puromycin.

Memory loss in mice

Long-term

synaptic plasticity In neuroscience, synaptic plasticity is the ability of synapses to Chemical synapse#Synaptic strength, strengthen or weaken over time, in response to increases or decreases in their activity. Since memory, memories are postulated to be represent ...

, such as is required for

memory Memory is the faculty of the mind by which data or information is encoded, stored, and retrieved when needed. It is the retention of information over time for the purpose of influencing future action. If past events could not be remembe ...

processes, requires morphological changes at protein level. As puromycin inhibits protein synthesis in eukaryotic cells, researchers were able to show that injections of this drug will result in both short-term as well as long-term memory loss in mice.

References

{{Protein synthesis inhibitor antibiotics Protein synthesis inhibitor antibiotics Nucleosides Carboxamides Eukaryotic selection compounds 4-Methoxyphenyl compounds Dimethylamino compounds