Metabolism of biomolecules/genetics material transmission

A       IDENTITAS

          Hari/Tanggal    : Kamis/ 26 Juli 2012

          Waktu               : 09.00-12.00

B       KEGIATAN                                        

         Metabolism of biomolecules/genetics material transmission   

C       RINGKASAN KEGIATAN

            Hari ini kami belajar tentang aliran informasi genetik, yang memebahas masalah DNA sampai terbentuk protein. Dosen pengajarnya seorang profesor, beliau mengajarkan masalah basic dari peristiwa transkripsi yang rumit, yang melibatkan banyak preotein dan enzim pada organisme prokariotik maupun eukariotik. Kami sempat mengunjungi laboratorium rekayasa genetik beliau. Beliau merekayasa Arabidopsys Thaliana dan Tobacco. Ternyata Arabidopsys adalah tanaman sejenis rumput yang memiliki sedikit sequen DNA yang telah  dipelajari dan diketahui sequences DNAnya, dimana hasil penelitian terhadap Arabidopsys dapat diaplikasikan pada tanamana agricultural yang lain

An overview of the flow of information through the cell

The flow of information in an eucaryotic cell:

Step 1: Selected site on the DNA are transcribed into pre-mRNA

Step 2: Pre-mRNA are processed into mRNA (messenger RNA)

Step 3: The mRNA are transported out of the nucleus

Step 4: mRNA are translated into polypeptides by ribosomes that move along the mRNA

 Genes can be expressed with different efficiencies

Gene A is transcribed and translated much more efficiently than gene B.

This allows the amount of protein A in the cell to be much greater than that

of protein B.

RNA is a polymer composed of alternating units of ribonucleotides

connected through a phosphodiester bond.

1.             Pentose (five carbon) sugar:

RNA contains ribose, which have a hydroxyl group on the 2’ carbon of the

ribose sugar

2.             Nitrogen bases: A,U,G,C

The base uracil in place of thymine

3.              Phosphate group

From DNA to RNA (transcription)

Transcription: The synthesis of an RNA from a DNA template.

Because its nucleotide sequence is complementary to that of the gene from which it is transcribed, the mRNA retains the same information as the gene it self.

The use of messenger RNA allows the cell to separate information storage from information utilization

While the gene remains stored away in the nucleus as part of a huge, stationary DNA molecule, its information can be imparted to a much smaller, mobile nucleic acid that pass into the cytoplasm. Once in the cytoplasma, the mRNA can serve as a template for synthesis of a large number of polypeptide chain.

Materials involved in transcription:

• Substrate: NTPs (ATP, UTP, GTP, CTP)

• Template: DNA

• DNA-dependent RNA polymerase: Enzyme that are able to incorporate nucleotides, one at a time, into a strand of RNA according to a DNA template. 1 in prokaryotes, 3 in eukaryotes.

• Other protein factors

General properties of DNA-dependent RNA polymerases

(i) Polarity: RNA polymerase reads the DNA template in the 3’→5’ direction while synthesizing RNA in the 5’→3’ direction

(ii) DNA template: Either strand of a DNA double helix can serve as a

template for RNA synthesis

RNA Polymerase direction is determined by the orientation of the promoter

sequence.

The promoter region in higher eukaryotes:

• The TATA box is located approximately 30 base pairs from the mRNA

start site.

• Two or more promoter-proximal elements are found 100 and 200 bp

upstream of the mRNA start site. The CCAAT box and the GC-rich box

are shown here.

• Other upstream elements include the sequences GCCACACCC and

ATGCAAAT.

Transcription Factors and RNA polymerases bind to the promoter region of a gene :

Three stages of RNA synthesis:

• Initiation:

RNA polymerase binds to the promoter sequence in duplex DNA, then melts duplex DNA near the transcription start site to form a transcription “bubble” before catalyzes the phosphodiester linkage of two initial NTPs.

• Elongation:

Polymerase advance the 3’ to 5’ of the template strand, melting the duplex DNA and adding NTPs to growing RNA.

• Termination:

Termination occurs when the RNA polymerase encounter a specific termination sequence.

Transcription in procaryotes:

- Initiation

- Elongation

- Termination

Differences between Prokaryotic and Eukaryotic Gene Expression

• Eukaryotic genes contain introns; prokaryotic genes do not.

• An eukaryotic mRNA code for 1 gene; A prokaryotic mRNA code for several related genes at one time.

• Eukaryotic mRNA must be moved to cytoplasm; prokaryotic mRNA is already in the cytoplasm and translation starts before transcription is complete.

Transcription initiation in eukaryotes requires many proteins

• RNA polymerase II is required for the synthesis of mRNA.

• Unlike the E. coli RNA polymerase holoenzyme, RNA polymerase II require a number of additional proteins called “general transcription factors” in order to specifically bind to a promoter and initiate transcription.

• Eukaryotic promoters are composed of a variety of different cis sequence elements which recruit some of these trans-acting factors through DNA-protein interactions.

• Protein-protein interactions also occur and account for many of the multi-component complexes found at eukaryotic promoters.

Eukaryotic RNA polymerases consist of many subunits

Post-transcriptional modifications:

In eukaryotic cells, a genetic process for the conversion of a primary transcript RNA to mature RNA

Processing of the human globin mRNA

1st Step: RNA capping

the 5’ end of the new RNA molecule is modified by addition of a cap that consists of a modified guanine nucleotide

The structure of the cap at the 5’ end of eucaryotic mRNA molecules

The methylguanosine cap at the 5’ end of an mRNA serves several functions:

1. It prevents the 5’ end of the mRNA from being digested by exonucleases.

2. It aids in transport of the mRNA out of the nucleus.

3. It plays an important role in the initiation of mRNA translation.

4. It signifies the 5’ end of eucaryotic mRNAs, and this landmark helps the cell to distinguish mRNAs from the other types of RNA molecules present in the cell.

2nd Step : RNA splicing

The intron sequences are removed from the newly synthesized RNA

Structure of two human genes showing the arrangement of exons and introns

The pre-RNA splicing reaction

1. a specific adenine nucleotide in the intron sequence (indicated in red) attacks the 5’ splice site and cuts the sugar phosphate backbone of the RNA at this point.

2. The cut 5’ end of the intron becomes covalently linked to the adenine nucleotide, thereby creating a loop in the RNA molecule.

3. The released free 3’-OH end of the exon sequence then reacts with the start of the next exon sequence, joining the two exons together and releasing the intron sequence in the shape of a lariat.

4. The two exon sequences become joined into a continuous coding sequence; the released intron sequence is eventually degraded.

The RNA splicing reaction is performed on the C-terminal domain of RNA polymerase II (CTD)

- key steps in RNA splicing are performed by RNA molecules rather than proteins

- These RNA molecules are relatively short (less than 200 bp each), and there are five of them (U1, U2, U4, U5, and U6) involved in the major form of pre-mRNA splicing, know as SnRNAs (small nuclear RNAs).

- Each SnRNA is complexed with at least seven protein subunits to form a snRNP (small nuclear ribonucleo protein). These snRNPs form the core of the spliceosome, the large assembly of RNA and protein molecules that performs pre-mRNA splicing in the cell.

3rd Step : RNA 3’ polyadenylation

Consensus nucleotide sequences that direct cleavage and polyadenylation to form 3’ end of a eucaryotic mRNA

Poly(A) tail invariably begins approximately 20 nucleotides downstream from the sequence AAUAAA, which serve as a recognition site for the assembly of a complex of proteins that carry out the processing reactions at the 3’ end of the mRNA.