User talk:Aravindabharathi/Alpha operon

The α operon in Escherichia coli contains genes coding for 4 ribosomal proteins (r-proteins) and also the rpoA gene, which codes for the α subunit of RNA polymerase. From a regulatory standpoint, the α operon is quite unique since control is mainly through translational repression by one of the gene products. However, the influence of this repression on the expression of rpoA is quite weak, and differential expression of the r-proteins and the α subunit can occur. Even though the expression of rpoA is differentially regulated to a very large extent, it is considered a part of the same operon since it is transcribed along with the other genes. It is the translation of the genes where the regulation occurs. Although the genes which code for r-proteins are divided into many different operons, all these genes are coordinately regulated, and all r-proteins are synthesised at the same rate.^[1]

Structure of the α operon

The α operon contains 5 structural genes, in the order listed^[2]:

rpsM, encoding the S13 r-protein
rpsK, encoding the S11 r-protein
rpsD, encoding the S4 r-protein
rpoA, which codes for the α subunit of RNA polymerase
rplQ, encoding the L17 r-protein

Order of genes on the α operon

Functional importance

Ribosomal proteins

S13 is one of the proteins which make up the 30S subunit of the bacterial ribosome.^[3] Along with the r-protein S12, it is thought to be responsible for controlling the movement of the mRNA:tRNA complex through the ribosome during protein synthesis.^[4]
S11 is found on the 30S ribosomal subunit and is vital for the selection of the correct tRNA corresponding to a codon during protein synthesis.^[5]
S4 is another protein on the 30S subunit. Apart from its functions on the ribosome, it functions as a tranlational repressor, preventing the translation of the mRNA sequence of the α operon when not associated into a ribosome. Also, it is an antitermination factor, and slows down termination during the transcription process. It can act as an antitermination factor since it is capable of binding to RNA polymerase.^[6]
L17 is found on the 50S subunit of the bacterial ribosome. Along with another r-protein L2, it can form a complex consisting of itself, L2, and a tRNA molecule which has a site for another tRNA molecule to bind to. In this way, it is vital for maintaining continuity during protein synthesis in the ribosome.^[7]

α subunit of RNA polymerase

RNA polymerase, also known as RNAP, is one of the most pivotal enzymes in the process of transcription. It is responsible for the de novo synthesis of the polynucleotide RNA molecule from a template DNA strand. RNA polymerase is a complex made up of five proteins, denoted as the α, β, β', ω, and σ subunits.

Two copies of the α subunit are present in one RNAP molecule, and are denoted as α^I and α^II. The α subunit is imperative for the assembly of RNAP from its constitutent proteins, which are present in different regions of the genome. The assembly reaction proceeds as follows:

2α → α₂ → α₂β → α₂ββ' → α₂ββ'σ

RNAP subunit α is also capable of directly binding to DNA. Some "strong" promoter have a supplementary promoter element upstream of the normal promoter site, in addition to the promoter itself. By recognising and binding to these promoters, RNAP subunit α enhances the stability of the binding of RNAP to the DNA, which results in an improvement in the efficiency of transcription initiation. ^[8]

Regulation

The α operon mRNA has a pseudoknot structure which stimulates the binding of the 30S subunit of the ribosome. It is, however, inactive at low temperatures and prevents the binding of the methionine-carrying tRNA, the first step in translation. It gets activated at higher temperatures, and translation can occur.^[9]

Translational repression of r-protein expression

In general, the regulation of r-protein synthesis occurs by post-translational feedback inhibition by "free" r-proteins which are not part of a ribosome. The repressive ability of the r-protein is removed when they are assembled together into a ribosome. This ensures that all r-proteins are synthesised at the same rate. In the α operon, the feedback inhibitor is the S4 protein, encoded by the gene rpsD.

The S4 protein is capable of binding to a single site on the mRNA synthesised from the α operon. By changing the shape of the mRNA, the binding of S4 prevents the formation of the mRNA-ribosome complex required for translation. Since the binding site of S4 is distinct from the Shine-Dalgarno sequence of the mRNA, this is a form of allosteric regulation.^[10]

Regulation of rpoA expression

Translational repression of the synthesis of the r-proteins occurs due to a change in the mRNA conformation caused by the binding of S4. However, the new conformation still permits the the translation of the mRNA from the rpoA gene, which codes for the α subunit of RNAP. This allows the differential expression of the rpoA gene with respect to its neighbours on the operon. Small changes to the rate of α subunit synthesis are observed in response to repression by S4, but this is attributed to a decrease in the stability of the mRNA caused by the binding of S4 to it. It is also predicted that the synthesis of the α subunit takes place from separate mRNA strands, which are independent of the α operon transcripts, and comprise solely of transcripts of the rpoA gene.

Eukaryotic analogues

Studies of the α subunit have shown that the amino acid sequence of the subunit in prokaryotes matches closely the sequence of the α subunit found in plant chloroplasts. This is one of the many supporting evidences for the endosymbiotic theory of the origin of organelles.

The α-subunit contains an amino-terminal domain (denoted αNTD) and a carboxy-terminal domain (denoted αCTD), which are independently folded. Large sequences of the αNTD have been found to be conserved across eukaryotic RNAP I, II and III. Primarily due to this, the role of the αNTD sequence in the assembly of the RNAP complex is believed to be conserved in eukaryotes and archaea also. However, the αCTD sequence from prokaryotes is not seen in archaea or eukaryotes, and so the promoter recognition functions of the α subunit are absent in those organisms.

References

^ D, Dean, and Nomura M. “Feedback Regulation of Ribosomal Protein Gene Expression in Escherichia Coli., Feedback Regulation of Ribosomal Protein Gene Expression in Escherichia Coli.” Proceedings of the National Academy of Sciences of the United States of America, Proceedings of the National Academy of Sciences of the United States of America 77, 77, no. 6, 6 (June 1980): 3590, 3590–3594. doi:10.1073/pnas.77.6.3590.
^ Thomas, M.S., D.M. Bedwell, and M. Nomura. “Regulation of α Operon Gene Expression in Escherichia Coli: A Novel Form of Translational Coupling.” Journal of Molecular Biology 196, no. 2 (July 20, 1987): 333–345. doi:10.1016/0022-2836(87)90694-2.
^ http://biocyc.org/ECOLI/NEW-IMAGE?type=ENZYME&object=EG10912-MONOMER
^ Cukras, Anthony R, Daniel R Southworth, Julie L Brunelle, Gloria M Culver, and Rachel Green. “Ribosomal Proteins S12 and S13 Function as Control Elements for Translocation of the mRNA:tRNA Complex.” Molecular Cell 12, no. 2 (August 2003): 321–328.
^ Nomura, Masayasu. “Bacterial Ribosome.” Bacteriological Reviews 34, no. 3 (September 1970): 228–277.
^ Torres, Martha, Ciarán Condon, Joan-Miquel Balada, Craig Squires, and Catherine L. Squires. “Ribosomal Protein S4 Is a Transcription Factor with Properties Remarkably Similar to NusA, a Protein Involved in Both Non-ribosomal and Ribosomal RNA Antitermination.” The EMBO Journal 20, no. 14 (July 16, 2001): 3811–3820. doi:10.1093/emboj/20.14.3811.
^ Metspalu, Ene, Toivo Maimets, Mart Ustav, and Richard Villems. “A Quaternary Complex Consisting of Two Molecules of tRNA and Ribosomal Proteins L2 and L17.” FEBS Letters 132, no. 1 (September 14, 1981): 105–108. doi:10.1016/0014-5793(81)80438-3.
^ Ebright, Richard H, and Steve Busby. “The Escherichia Coli RNA Polymerase α Subunit: Structure and Function.” Current Opinion in Genetics & Development 5, no. 2 (April 1995): 197–203. doi:10.1016/0959-437X(95)80008-5.
^ Spedding, Gary, Thomas C. Gluick, and David E. Draper. “Ribosome Initiation Complex Formation with the Pseudoknotted α Operon Messenger RNA.” Journal of Molecular Biology 229, no. 3 (February 5, 1993): 609–622. doi:10.1006/jmbi.1993.1067.
^ Spedding, G., and D. E. Draper. “Allosteric Mechanism for Translational Repression in the Escherichia Coli Alpha Operon.” Proceedings of the National Academy of Sciences 90, no. 10 (May 15, 1993): 4399–4403. doi:10.1073/pnas.90.10.4399.

[1] D, Dean, and Nomura M. “Feedback Regulation of Ribosomal Protein Gene Expression in Escherichia Coli., Feedback Regulation of Ribosomal Protein Gene Expression in Escherichia Coli.” Proceedings of the National Academy of Sciences of the United States of America, Proceedings of the National Academy of Sciences of the United States of America 77, 77, no. 6, 6 (June 1980): 3590, 3590–3594. doi:10.1073/pnas.77.6.3590.

[2] Thomas, M.S., D.M. Bedwell, and M. Nomura. “Regulation of α Operon Gene Expression in Escherichia Coli: A Novel Form of Translational Coupling.” Journal of Molecular Biology 196, no. 2 (July 20, 1987): 333–345. doi:10.1016/0022-2836(87)90694-2.

[3] ttp://biocyc.org/ECOLI/NEW-IMAGE?type=ENZYME&object=EG10912-MONOMER

[4] Cukras, Anthony R, Daniel R Southworth, Julie L Brunelle, Gloria M Culver, and Rachel Green. “Ribosomal Proteins S12 and S13 Function as Control Elements for Translocation of the mRNA:tRNA Complex.” Molecular Cell 12, no. 2 (August 2003): 321–328.

[5] Nomura, Masayasu. “Bacterial Ribosome.” Bacteriological Reviews 34, no. 3 (September 1970): 228–277.

[6] Torres, Martha, Ciarán Condon, Joan-Miquel Balada, Craig Squires, and Catherine L. Squires. “Ribosomal Protein S4 Is a Transcription Factor with Properties Remarkably Similar to NusA, a Protein Involved in Both Non-ribosomal and Ribosomal RNA Antitermination.” The EMBO Journal 20, no. 14 (July 16, 2001): 3811–3820. doi:10.1093/emboj/20.14.3811.

[7] Metspalu, Ene, Toivo Maimets, Mart Ustav, and Richard Villems. “A Quaternary Complex Consisting of Two Molecules of tRNA and Ribosomal Proteins L2 and L17.” FEBS Letters 132, no. 1 (September 14, 1981): 105–108. doi:10.1016/0014-5793(81)80438-3.

[8] Ebright, Richard H, and Steve Busby. “The Escherichia Coli RNA Polymerase α Subunit: Structure and Function.” Current Opinion in Genetics & Development 5, no. 2 (April 1995): 197–203. doi:10.1016/0959-437X(95)80008-5.

[9] Spedding, Gary, Thomas C. Gluick, and David E. Draper. “Ribosome Initiation Complex Formation with the Pseudoknotted α Operon Messenger RNA.” Journal of Molecular Biology 229, no. 3 (February 5, 1993): 609–622. doi:10.1006/jmbi.1993.1067.

[10] Spedding, G., and D. E. Draper. “Allosteric Mechanism for Translational Repression in the Escherichia Coli Alpha Operon.” Proceedings of the National Academy of Sciences 90, no. 10 (May 15, 1993): 4399–4403. doi:10.1073/pnas.90.10.4399.

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