Protein synthesis.

There provided best content for learning that how to PROTEIN synthesis and how the proteins are sorting to different organelles.

1. UTTAM MEMORIAL COLLEGE PATELPALI RAIGARH (C.G.) SUB- MOLECULAR CELL BIOLOGY TOPIC – PROTEIN SYNTHESIS IN FREE & BOUND POLYSOMES & UPTAKE INTO E.R. GUIDED BY-. SUBMITTED BY – MISS SUSHMITA MA’AM KRISHN KUMAR SAHU (MOLECULAR BIO. TEACHER) (MSc. ZOOLOGY 2 ND SEM)

2. Introduction. Transcription.
transcription Machinery. Transcription in prokaryotes.
Transcription in Eukaryotes. Mechanism of Transcription Genetic code. Translation.
Translation Machinery.
Mechanism of Translation. Uptake to ER (sorting & transporting). Conclusion.
Reference. SYNOPSIS:-

3. 1. INTRODUCTION:- Protein synthesis(translation) is the production of a polymer of a chain of amino acids which produces a functioning protein. Ribosomes are the structures that synthesize the protein chain. Protein synthesis requires a tRNA (transfer RNA) which helps in the transfer of an amino acid molecule to the growing chain. tRNA has anti-codons for the codons present on the mRNA molecule which helps to recognize the place of single amino acid. POLYSOME :- polyribosome (or polysome or ergosome ) is a group of ribosomes translating to mRNA into protein. It consists of a complex of an mRNA molecule and two or more ribosomes that act to translate mRNA instructions into polypeptides.

4. 2. TRANSCRIPTION:- It is process of copying information from template strand to DNA into RNA Mediated by RNA polymerase. Take place in the Nucleus of Eukaryotic cells. Converting process of DNA Into RNA is called transcription process..

5. Strand of DNA:- Two type of strand are present.
1. Template Strand:- Strand of DNA with 3 to 5 polarity that direct synthesis of RNA is called template strand.
2. coding strand:- DNA strand with 5 to 3 polality that Strand Are sensing strands.

6. Transcription Unite: - The segment of DNA . Between the sites of Initiation and termination of transcription the place known as a Transcription unite. Contains 3 Rigion 1. Promotar Rigion . 2. Structural gene Rigion . 3 Terminal Rigion ..

7. 1. PROMOTAR :- It is a region on DNA molecule to which an RNA polymerase binds and initiates transcription. Promoter is located upstream (to the left) of the structural gene, inwards 5’ end of coding strand, 3’ end of template strand.

8. 2. STRUCTURAL GENE:- A gene that codes for the amino acid sequence of a protein (such as an enzyme) or for a ribosomal RNA or transfer RNA Structural gene is a gene that codes for any RNA or protein product other than a regulatory factor (i.e. regulatory protein).

9. 3. TRANSCRIPTION MACHINERY:- Enzyme RNA Polymerase
DNA template
Four types of ribonucleotides triphosphates (ATP, CTP, GTP and UTP)
Divalent metal ions Mg2+ or Mn2+ as a cofactor.

10. 1. Enzyme RNA polymerase:- It is a large complex consisting of 4 subunits which make the core enzyme and Sigma subunit and p (Rho) subunit. 1. Core Enzyme :-It has 4 subunits.
2. Sigma Subunit :- it Recognises promoter and initiates synthesis.
3. Rho Subunit :-Helps in termination of transcription .& inhibit transcript Eukaryotes contains 3 types of Polymerase but prokaryotes has single. RNA Polymerase I :- Synthesizes rRNA RNA Polymerase II :- Sythesizes mRNA
RNA Polymerase III:- Synthesizes tRNA

11. 4. TRANSCRIPTION MECHANISM:- 1. Prokaryotes:- Transcription In prokaryotes the structural genes are polycistronic and continuous The process of transcription is completed in following steps: 1. Initiation. 2. Elongation. 3. Termination.

12. 1. INITIATION:- RNA polymerase reaches the promoter region and binds to it. It has a sigma factor (also called initiation factor). The enzyme recognizes the promoter by its sigma factor RNA polymerase initiates transcription.
It uses nucleoside triphosphates as substrate and polymerases in a template depended fashion following the rule of complementarity.
It also facilitates opening of helix.

13. 2. ELONGATION:- Elongation is the stage when the RNA strand gets longer, thanks to the addition of new nucleotides. During elongation, RNA polymerase “walks ” along one strand of DNA, known as the template strand, in the 3’ to 5’ direction. For each nucleotide in the template, RNA polymerase adds a matching (complementary) RNA nucleotide to the 3’ end of the RNA strand.

14. 3. TERMINATION:- When RNA polymerase reaches terminator region a specific chain terminating protein called rho (p) factor (also called termination factor) stops the synthesis of RNA chain.
It separates RNA polymerase as well as the newly formed RNA strand. RNA and RNA polymerase fall off and it results in termination of transcription.

15. 2. Transcription in Eukaryotes:- It is more complex than that of prokaryotic transcription. It occurs inside nucleus and two types of semi-autonomous cell organelles (e.g.. Mitochondria and plastids).
It occurs in G, and G₂ phases of cell cycle.

16. Transcriptional unit has only one gene ( monocistronic )
Initiation requires proteins called transcription factors.
Sigma factor is absent No primer is required.
RNAs are released and processed in the nucleus.
Coupled transcription-translation is not possible.
Greater part of products pass from nucleus into cytoplasm. SPECIAL FEATURE OF TRANSCRIPT. (EUKARYOTES)

17. Processing of Primary Transcript:- The nascent (newly formed) RNA synthesized by RNA polymerase II is called hnRNA or primary transcript. The primary transcript contains: Unwanted base sequences ( introns ). Useful base sequences ( exons ). Four steps are For primary transcript:- 1. Capping 2. Tailing 3. Splicing 4. Ligation

18. 1. Capping at 5’ end:- It is addition of a cap to 5' end of hnRNA . It occurs shortly after the start of transcription. The cap contains an unusual nucleotide ig . Methyl guanosine triphosphate ( mGppp ) 2. Tailing at 3’ end:- It is addition of poly A tail at 3’ end of hn mRNA with the help of poly A polymerase.
The poly A tail contains adenylate residues (about 200-300).
Polyadenylation protects the 3’ end from degradation by exonucleases.

19. Hr RNA Ligation

20. 3. Splicing:- The process by which introns, the noncoding regions of genes, are excised out of the primary messenger RNA transcript, and the exons (i.e., coding regions) are joined together to generate mature messenger RNA.

21. 4. Ligation:- The ends of the adjacent exons are joined together by ligase enzyme. The snRNPs are formed by association of proteins with small nuclear RNAS. ully processed hnRNA is now called mRNA and is released from the nucleus into the cytoplasm for translation.

22. Types of RNA:-

23. 5. Genetic code:- The genetic code is a set of three-letter combinations of nucleotides called codons, each of which corresponds to a specific amino acid Genetic code is a triplet. There are 64 codes, out of which 61 codons code for amino acids But 3 codon do not code the amino acid.
The codon is unambiguous and specific i.e. One codon codes for only one amino acids. Some amino acids are coded by more than one codon, hence the code is degenerate.

24. Stop codon

25. 6. TRANSLATION:- Translation is the process in which ribosomes in the cytoplasm or endoplasmic reticulum synthesize proteins after the process of transcription of DNA to RNA in the cell’s nucleus. Called translation. The entire process is called gene expression.

26. 7. Machinery of translation:-

27. Ribosomal sites:-

28. 8. Translation Mechanism :-

29. 1.Activation of Amino Acids In the presence of ATP, an amino acid combines with its specific aminoacyl- tRNA synthetase .
Mg2+ is required. Activation is the covalent coupling of amino acids to specific adapter molecules. The adapter molecules are called transfer RNA ( tRNA ). There is at least on tRNA for each of the 20 naturally occurring amino acids.

30. 2. Charging or Aminoacylation of tRNA The process of attaching an amino acid to its corresponding transfer RNA ( tRNA ) is called amino acid activation, also known as tRNA charging or aminoacylation . complex. Enzyme and AMP are released. tRNA complexed with amino acid is called charged tRNA The amino acid is linked to 3-OH-end of tRNA through its COOH group

31. 3. Initiation:- mRNA attaches itself to smaller subunit of ribosome in the region of its cap. Cap has nucleotides complementary to the nucleotides present at 3’ end of rRNA . Attachment is such that initiation codon of mRNA comes to lie at P-site. Attachment of large unite of ribosomes.

32. 3.Elongation(Polypeptide Chain Formation): Elongation (Polypeptide Chain Formation)
An aminoacyl tRNA complex reaches the A-site and attaches to mRNA codon next to initiation cotton with the help of its anticodon.
A peptide bond is established between the carboxyl group (-COOH) of amino acid attached to tRNA at P-site and amino group (-NH2) of amino acid attached to tRNA at A-site.

33. 4. Termination:- Polypeptide synthesis is terminated when a nonsense codon of mRNA reaches the A- site. There are three nonsense codons:
UAA (ochre) UAG (amber) UGA (opal). Nonsense codons are not recognised by any of the tRNAs .
Therefore, no more aminoacyl tRNA reaches the A-site.
The P-site tRNA is hydrolysed and the completed polypeptide is released in presence of GTP-dependent release factor.
Ribosome moves over the nonsense codon and slips off the mRNA codon
The two subunits of ribosome separate or undergo dissociation in the
dissociation factor (DF).

34. Proteins synthesis in bound polyribosome:- Membrane-bound ribosomes, attached to the cytosolic side of the ER membrane, are engaged in the synthesis of proteins that are being concurrently translocated into the ER. Free ribosomes, unattached to any membrane, synthesize all other proteins encoded by the nuclear genome. Membrane-bound and free ribosomes are structurally and functionally identical. They differ only in the proteins they are making at any given time. When a ribosome happens to be making a protein with an ER signal sequence, the signal directs the ribosome to the ER membrane.

35. Cont.. Since many ribosomes can bind to a single mRNA molecule; a polyribosome is usually formed, which becomes attached to the ER membrane, directed there by the signal sequences on multiple growing polypeptide chains . The individual ribosomes associated with such an mRNA molecule can return to the cytosol when they finish translation near the 3’ end of the mRNA molecule. The mRNA itself, however, remains attached to the ER membrane by a changing population of ribosomes, each transiently held at the membrane by the translocator . In contrast, if an mRNA molecule encodes a protein that lacks an ER signal sequence, the polyribosome that forms remains free in the cytosol, and its protein product is discharged there. Therefore, only those mRNA molecules that encode proteins with an ER signal sequence bind to rough ER membranes.

36. 10 Protein uptake to ER/Sorting/ Targetting :- Protein targeting or protein sorting is the biological mechanism by which proteins are transported to their appropriate destinations within or outside the cell. Process:- 1. Co translation translocation. . 2. Post translation translocation. 1. CO TRANSLATION TRANSLOCATION:- RER Ribosomes. Secretory protein. ER, GB, LYSOSOME and PLASMA MEMBRANE

37. Mechanism:- Protein which is forming in translation process,are taken with ribosome Ñ terminal of protein have 6- 12 hydrophobic amino acid. This n terminal contains signals sequence. Cytoplasm of cells contains SRP ( signal recognise protein). This SRP molecule binds to the signal sequence and inhibit the translation process for some time. Pm of any other cell contains SRP BINDING RECEPTOR. & Specific channel called TRANSLOCON , this TRANSLOCON are always in inactive form unless any SRP did not attach into SRP receptor. When SRP binds to the it receptor, TRANSLOCON gonna activated.

38. Cont.. After binding,it activates the channel and open the channel,and protein enter into the ER. Signal peptidase cut the signal sequence and start the folding process. Then Ribosome and SRP protein are removed in the cytoplasm.

39. Post translation translocation:- 1. In prokaryotes:- Translated protein are present in cytoplasm of bacteria. SEC-A Protein are present in cytoplasm. Transmembraned protein SEC- Y present in pm. Sec-A binds to the Sec-Y protein. Then shortly shortly protein entered into other .

40. Post translation translocation:- 2. In Eukaryotes:- fully translated protein are present. C terminal of protein binds chaperon protein And N terminal no need to add SRP PROTEIN. It attached on ER. MEMBRANE where present SEC- 61 protein. And after attachment it activates the channel and n terminal entered into ER. then peptidase enzymes cuts the protein and folding start.

41. 11. Conclusion:- Translation is the process in which a polypeptide chain obtained of different amino acids from the base sequence of an mRNA molecule in association with a ribosome. In other word “it is process by which genetic information present in mRNA are translated into the language of Protein” Translation is much more complex than transcription as it involves initiation clongation and termination of polypeptide chains. 12. Reference:- HUMAN PHYSIOLOGY BOOK BY DR. Anant prakash gupta And TAKEN FROM FAMOUS NEET UNACADEMY APP . (TEACHER DR. SACHIN KAPOOR AND RITU RATTEWAL MAAM)