PROTIEN SYNTHESIS

PROTEIN SYNTHESIS

            In 1958 Crick proposed that DNA determines the sequence of amino acid in a polypeptide through mRNA, this is the main principle or central dogma of protein synthesis which involves transcription and translation.

                                  

Introduction –

1.    Protein bio-synthesis is the process in which cells build or manufacture proteins.

2.    The term is sometimes used to refer only to protein translation but more often it refers to a multi-step process, beginning with amino acid synthesis and transcription of nuclear DNA into m RNA, which is then used as input for translation.

3.    The cistron DNA is transcribed into a variety of RNA intermediates.

4.    The last version is used as a template in synthesis of a polypeptide chain.

5.    Proteins can often be synthesized directly from genes by translating mRNA. When a protein must be available on short notice or in large quantities, a protein precursor is produced.

6.    A pro-protein is an inactive protein containing one or more inhibitory peptides that can be activated when the inhibitory sequence is removed by proteolysis during posttranslational modification.

7.    A pre-protein is a form that contains a signal sequence (an N-terminal signal peptide) that specifies its insertion into or through membranes, i.e.targets them for secretion.

8.     The signal peptide is cleaved off in the endoplasmic reticulum.

9.    For synthesis of protein, a succession of t-RNA molecules, amino acids, mRNA molecule and ribosome (ribosomal RNA and more than 50 different proteins).

Transcription -                         

1.     In transcription an mRNA chain is generated, with one strand of the DNA double helix in the genome as template. This strand is called the template strand.

2.     Transcription can be divided into 3 stages: Initiation, Elongation, and Termination.

3.     Each regulated by a large number of proteins such as transcription factors and coactivators that ensure that the correct gene is transcribed.

4.     The DNA strand is read in the 3' to 5' direction and the mRNA is transcribed in the 5' to 3' direction by the RNA polymerase (RNAP).

5.     Transcription occurs in the cell nucleus, where the DNA is held. The DNA structure of the cell is made up of two helixes made up of sugar and phosphate held together by the bases.

6.     The sugar and the phosphate are joined together by covalent bond. The DNA is "unzipped" by the enzyme helicase.

7.     Leaving the single nucleotide chain open to be copied.

8.     RNA polymerase reads the DNA strand from 3-prime (3') end to the 5-prime (5') end, while it synthesizes a single strand of messenger RNA in the 5'-to-3' direction.

9.     The general RNA structure is very similar to the DNA structure, but in RNA the nucleotide uracil takes the place that thymine occupies in DNA.

10.  The single strand of mRNA leaves the nucleus through nuclear pores, and migrates into the cytoplasm.

11.  The first product of transcription differs in prokaryotic cells from that of eukaryotic cells, as  in  prokaryotic  cells the  product  is  mRNA,  which  needs  no  post-transcriptional modification, whereas, in eukaryotic cells, the first product is called primary transcript, that needs post-transcriptional modification (capping with 7-methyl-guanosine, tailing with a poly A tail) to give hnRNA (heterophil nuclear RNA).

12.  hnRNA then undergoes splicing of introns (noncoding parts of the gene) via spliceosomes to produce the final mRNA.

Translation –

1.       The synthesis of proteins is known as translation.

2.       Translation occurs in the cytoplasm, where the ribosomes are located.

3.       Ribosomes are made of a small and large subunit that surround the mRNA.

4.       In translation, mRNA  is decoded to produce a specific polypeptide according to the rules specified by the trinucleotide genetic code.

5.       This uses an mRNA sequence as a template to guide the synthesis of a chain of amino acids that form a protein.

6.       Translation proceeds in four phases: activation, initiation, elongation, and termination (all describing the growth of the amino acid chain, or polypeptide that is the product of translation).

7.        In activation, the correct amino acid (AA) is joined to the correct transfer RNA (tRNA).

8.        The AA is joined by its carboxyl group to the 3' OH of the tRNA by an ester bond.

9.        When the tRNA has an amino acid linked to it, it is termed "charged". Initiation involves the small subunit of the ribosome binding to 5' end of mRNA with the help of initiation factors (eIF₁, eIF_2, eIF₃), other GTP that assist the process.

10.    eIF₁ and eIF₃ bind to a free 40s subunit. This helps to prevent a large subunits binding to it without an mRNA molecule and forming an inactive ribosome.

11.    eIF₂ complexed with GTP then binds to small subunit. It will assist the charged initiator tRNA to bind.

12.    The small subunit first binds to mRNA then activated tRNA with amino acid attached on start codon at P-site. Now large subunit attached on mRNA.

13.    Elongation occurs when the next aminoacyl-tRNA (charged tRNA) in line binds to the ribosome along with GTP and an elongation factor (eEFα, eEFβγ, eEF2).

14.    Termination of the polypeptide happens when the A site of the ribosome faces a stop codon (UAA, UAG, or UGA). When this happens, no tRNA can recognize it, but releasing factor (eRF) can recognize nonsense codons and causes the release of the polypeptide chain.

15.    The capacity of disabling or inhibiting translation in protein biosynthesis is used by some antibiotics such as anisomycin, cycloheximide, chloramphenicol, tetracycline, streptomycin, erythromycin, puromycin, etc.