Blasticidin S is an antibiotic that is used in biology research for selecting cells in cell culture. Cells of interest can express the blasticidin resistance genes BSD or bsr, and can then survive treatment with the antibiotic. Blasticidin S is a

nucleoside analogue Nucleoside analogues are nucleosides which contain a nucleic acid analogue and a sugar. Nucleotide analogs are nucleotides which contain a nucleic acid analogue, a sugar, and a phosphate group with one to three phosphates. Nucleoside and nucl ...

antibiotic, resembling the nucleoside

cytidine Cytidine (symbol C or Cyd) is a nucleoside molecule that is formed when cytosine is attached to a ribose ring (also known as a ribofuranose) via a β-N1- glycosidic bond. Cytidine is a component of RNA. It is a white water-soluble solid. which ...

. Blasticidin works against human cells, fungi, and bacteria, all by disrupting protein translation. It was originally described by Japanese researchers in the 1950s seeking antibiotics for rice blast fungus.

Chemistry

A nucleoside analog, blasticidin S resembles the nucleoside

. The chemical structure consists of a

cytosine Cytosine () ( symbol C or Cyt) is one of the four nucleobases found in DNA and RNA, along with adenine, guanine, and thymine (uracil in RNA). It is a pyrimidine derivative, with a heterocyclic aromatic ring and two substituents attached (an ...

molecule, linked to a glucuronic acid-derived ring, linked in turn to the

peptide Peptides (, ) are short chains of amino acids linked by peptide bonds. Long chains of amino acids are called proteins. Chains of fewer than twenty amino acids are called oligopeptides, and include dipeptides, tripeptides, and tetrapeptides. ...

N-methyl β-arginine.

Uses

Blasticidin S is widely used in cell culture for selecting and maintaining genetically manipulated cells. Cells of interest express the blasticidin S resistance genes BSD or bsr, and can then survive blasticidin S being added to the

culture media A growth medium or culture medium is a solid, liquid, or semi-solid designed to support the growth of a population of microorganisms or cells via the process of cell proliferation or small plants like the moss ''Physcomitrella patens''. Different ...

. Blasticidin S is typically used at 2–300

microgram In the metric system, a microgram or microgramme is a unit of mass equal to one millionth () of a gram. The unit symbol is μg according to the International System of Units (SI); the recommended symbol in the United States and United Kingdom whe ...

s per milliliter of media, depending on the type of cell being grown.

Mechanism of action

Blasticidin prevents the growth of both

eukaryotic Eukaryotes () are organisms whose Cell (biology), cells have a cell nucleus, nucleus. All animals, plants, fungi, and many unicellular organisms, are Eukaryotes. They belong to the group of organisms Eukaryota or Eukarya, which is one of the ...

and

prokaryotic A prokaryote () is a single-celled organism that lacks a nucleus and other membrane-bound organelles. The word ''prokaryote'' comes from the Greek πρό (, 'before') and κάρυον (, 'nut' or 'kernel').Campbell, N. "Biology:Concepts & Connec ...

cells. It works by inhibiting termination step of translation and peptide bond formation (to lesser extent) by the ribosome. This means that cells can no longer produce new proteins through

translation Translation is the communication of the meaning of a source-language text by means of an equivalent target-language text. The English language draws a terminological distinction (which does not exist in every language) between ''transla ...

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 synthesizing a protein. mRNA is created during the ...

. It is competitive with

puromycin Puromycin is an antibiotic protein synthesis inhibitor which causes premature chain termination during translation. Inhibition of translation Puromycin is an aminonucleoside antibiotic, derived from the '' Streptomyces alboniger'' bacterium ...

suggesting a highly similar binding site.

Biosynthesis

The first step in blasticidin S biosynthesis is the combination of

UDP-glucuronic acid UDP-glucuronic acid is a sugar used in the creation of polysaccharides and is an intermediate in the biosynthesis of ascorbic acid (except in primates and guinea pigs). It is made from UDP-glucose by UDP-glucose 6-dehydrogenase (EC 1.1.1.22) usin ...

with cytosine to form cytosylglucuronic acid (CGA). Given the product name, the enzyme that performs this combination is called

CGA synthase CGA may refer to: Companies and organizations * Canadian Gemmological Association * Coast Guard Administration of Taiwan * Controller General of Accounts of Pakistan * Central of Georgia Railway, in 1833, a.k.a. Central Rail Road and Banking C ...

. Cosmid cloning experiments from the Blasticidin S producer ''

Streptomyces griseochromogenes ''Streptomyces griseochromogenes'' is a bacterium species from the genus of ''Streptomyces'' which has been isolated from soil.Deutsche Sammlung von Mikroorganismen und Zellkulturenbr>/ref> ''Streptomyces griseochromogenes'' produces blasticidi ...

'', followed by evaluation of the putative biosynthetic gene cluster via heterologous reconstitution of Blasticidin S production in ''

Streptomyces lividans Snapalysin (, ''small neutral protease'', ''SnpA gene product (Streptomyces lividans)'') is an enzyme. This enzyme catalyses the following chemical reaction : Hydrolyses Hydrolysis (; ) is any chemical reaction in which a molecule of water b ...

'', indicated that a 20 Kbp gene cluster with 19 genes, plus possibly a peptidase outside the gene cluster that acts on the final leucylblasticidin S (LBS) intermediate, was sufficient for reconstitution of Blasticidin S biosynthesis.

Resistance genes

Resistance to blasticidin S can be conferred by either of two deaminases: BSD, originally isolated from ''

Aspergillus terreus ''Aspergillus terreus'', also known as ''Aspergillus terrestris'', is a fungus (mold) found worldwide in soil. Although thought to be strictly asexual until recently, ''A. terreus'' is now known to be capable of sexual reproduction. This saprotr ...

'' or bsr, isolated from ''

Bacillus cereus ''Bacillus cereus'' is a Gram-positive rod-shaped bacterium commonly found in soil, food, and marine sponges. The specific name, ''cereus'', meaning "waxy" in Latin, refers to the appearance of colonies grown on blood agar. Some strains are har ...

''. Both deaminases work by modifying blasticidin S directly, replacing the amine on the cytosine ring with a hydroxyl group, resulting in the inactive deaminohydroxy-blasticin S. bsr and BSD are the most commonly used resistance genes. The proteins produced from these genes enable the cells carrying them to produce proteins in the presence of blasticidin.

History

In the 1950s, a drug screening program was designed in Japan to discover a new antibiotic that prevents

blast disease ''Magnaporthe grisea'', also known as rice blast fungus, rice rotten neck, rice seedling blight, blast of rice, oval leaf spot of graminea, pitting disease, ryegrass blast, Johnson spot, neck blast, wheat blast, and Imochi (Japanese:稲熱) is a ...

by the

fungus A fungus ( : fungi or funguses) is any member of the group of eukaryotic organisms that includes microorganisms such as yeasts and molds, as well as the more familiar mushrooms. These organisms are classified as a kingdom, separately from t ...

'' Magnaporthe grisea''.Natural Products Isolation: Separation Methods for Antimicrobials, Antivirals, and Enzyme Inhibitors. Wagman G. H., Elsevier R. C.; p. 191 (1988).

References

{{Reflist

External links

Taxonomy of Streptomyces griseochromogenes
Protein synthesis inhibitor antibiotics Genetic engineering Pyrimidones Guanidines Eukaryotic selection compounds