Download - NUCLEIC ACIDS DNA, RNA & Protein Synthesis SDK September 24, 2013

NUCLEIC ACIDSDNA, RNA &

Protein SynthesisSDK

September 24, 2013

Road Map

1. What is nucleic Acid1. Types of nucleic acid2. Structural and functional characeristics of nucleic acid3. Nomenclature of Nucleic Acid Components

2. DNA1. Definition2. Structure of DNA3. Types of DNA4. DNA & Chromosomes

3. RNA1. Definition2. Structure& function of RNA

4. Difference between DNA & RNA5. Protein and Gene Expression

1. Protein Synthesis2. Transcription3. DNA Intones and Exons4. Translation5. Polypeptide chain6. Termination codon7. Summary

6. Brain Work

Nucleic Acids• Are the organic molecules derived from the

nucleus.

Friedrich Miescher in 1869• Isolated what he called nuclein from the nuclei of pus

cells• Nuclein was shown to have acidic properties, hence it

became called nucleic acid

Types of Nucleic Acid

• Deoxyribonucleic acid (DNA)• Ribonucleic acid (RNA)

The Distribution of Nucleic Acids in The Human Cell

• DNA is found in the nucleus with small amounts in mitochondria

• RNA is found throughout the cell

Nucleic Acid Structure

• Nucleic acids are polynucleotides

• Their building blocks are nucleotides


1. Primary- Basic component of NA

2. Secondary- Nucleoside and nucleotides

3. Tertiary- Formation of poly nucleotide chain

4. Quaternary- Pairing of two poly nucleotide

chains

Basic structure of Purine & Pyrimidine

Pyrimidines

Purines


O

N

N

NH2

O

CH2OPO

O-O-

OH

O

N

N

NH2

CH2OPO

O-

OH

O

N

N

AMP

CMP

3

5

Nomenclature of Nucleic Acid Components

Base Nucleoside Nucleotide Nucleicacid

PurinesAdenine Adenosine Adenylate RNA

Deoxyadenosine Deoxyadenylate DNA-------------------------------------------------------------------------------------Guanine Guanosine Guanylate RNA

Deoxy guanosine Deoxyguanylate DNA------------------------------------------------------------------------------------- PyrimidinesCytosine Cytidine Cytidylate RNA

Deoxycytidine Deoxycytidylate DNA-------------------------------------------------------------------------------------Thymine Thymidine Thymidylate DNA-------------------------------------------------------------------------------------Uracil Uridine Uridylate RNA

5-Fluorouracil 6-Mercaptopurine

Anticancer agents

Azidothymidine Dideoxyinosine

Antiretroviral agents

Nucleoside and base analogs can be used as anti-cancer and anti-virus drugs

Double Helix of DNA• DNA, or deoxyribonucleic acid, is the hereditary material in

humans and almost all other organisms.

• Nearly every cell in a person’s body has the same DNA. • Most DNA is located in the cell nucleus (where it is called

nuclear DNA), but a small amount of DNA can also be found in the mitochondria (where it is called mitochondrial DNA or mtDNA).

• Human DNA consists of about 3 billion bases, and more than 99 percent of those bases are the same in all people.

Why the people are different from each other

The order, or sequence, of these bases determines the information available for building and maintaining an organism, similar to the way in which letters of the alphabet appear in a certain order to form words and sentences.

• DNA contains two strands of nucleotides• H bonds hold the two strands in a double-helix

structure• A helix structure is like a spiral stair case• Bases are always paired as A–T and G-C• Thus the bases along one strand complement the

bases along the other

Traditionally, a DNA sequence is drawn from 5’ to 3’ end.

A shorthand notation for this sequence is ACGTA

The primary structure of DNA is the sequence

The 2ndry structure of DNA is a Double Helix of DNA

DNA TypesProperty A-DNA B-DNA Z-DNAHelix Handedness Right Right Left

Base Pairs per turn 11 10.4 12

Rise per base pair along axis

0.23nm 0.34nm 0.38nm

Pitch 2.46nm 3.40nm 4.56nm

Diameter 2.55nm 2.37nm 1.84nm

Width Widest Intermediate Narrowest

Conformation of Glycosidic bond

anti anti Alternating anti and syn

Major Groove Present Present Absent

Minor Groove Present Present Deep cleft

Helix : right handedBase pairs: almost perpendicular to the helix axis; 3.4 Å apartOne turn of the helix: 36 Å; ~10.4 base pairs Minor groove: 12 Å acrossMajor groove: 22 Å across

Normally DNA is B-form DNA

In eukaryotic cells,

DNA is folded into chromatin

DNA Tertiary Structure

DNA double helical structure coils round histones.

DNA bound to histones forms nucleosomes (10nm fibres)

Nucleosomes contain 146 nucleotidesNucleosomes

Repeating globular subunits of chromatin that consist of a complex of DNA and histone.

Compaction of DNA in a eukaryotic Chromosome

Supercoil = coil over coil

Functions of DNA and summary of structure

DNA consists of four bases—A, G, C, and T—that are held in linear array by phosphodiester bonds through the 3' and 5' positions of adjacent deoxyribose .

DNA is organized into two strands by the pairing of bases A to T and G to C on complementary strands. These strands form a double helix around a central axis.

The 3 x 109 base pairs of DNA in humans are organized into the haploid complement of 23 chromosomes.

Ribonucleic acid (RNA)

Ribonucleic acid (RNA)• Definition: RNA is polymer of ribonucleotides held

together by 3′,5′-phosphodiester bonds.

• The RNA is made from four types of ribonucleotides, ATP,GTP,CTP and UTP.

• These ribonucleotides are made from: Nitrogenous bases ; Adenine, Guanine, Uracil and Cytosine.Sugar –ribosePhosphate group

• Single stranded but usually forms intra-molecular base

pairs

• major and minor grooves are less pronounced

• Uracil instead of thymine

• Structural, adaptor and transfer roles of RNA are all

involved in decoding the information carried by DNA

RNA Structure

Type of RNA

Short name Functions

Messenger RNA mRNA Transfers genetic information from genes to ribosomes for synthesis of proteins

Transfer RNA tRNA Acts as an adaptors between mRNA and amino acids in protein synthesis

Ribosomal RNA rRNA •Provides structural framework for ribosomes. •Also helps in peptide bond formation (as an enzyme)

Types and functions of RNA Three major types:

Messenger RNA: 5-10%

Transfer RNA : 10-20%

Ribosomal RNA: 50-80%

Minor Types of RNA

Heterogeneous nuclear RNA

hnRNA Serves as precursor for mRNA and rRNA

Small nuclear RNA snRNA Involved in mRNA processing

Small nucleolar RNA

snoRNA Plays a key role in the processing of rRNA molecules.

Small cytoplasmic RNA

scRNA Involved in the selection of proteins for export

Micro RNA miRNA Involved in translation and mRNA degradation

Non-coding RNA ncRNA Involved in telomere synthesis, protein transport , tRNA processing etc

Messenger RNA (m-RNA)

Comprises only 5% of the RNA in the cellMost heterogeneous in size and base sequenceAll members of the class function as messengers carrying the information in a gene to the protein synthesizing machinery

Structural Characteristics of m-RNA

The 5’ terminal end is capped by 7- methyl guanosine triphosphate cap.

The cap is involved in the recognition of mRNA by the translating machinery

It stabilizes m RNA by protecting it from 5’ exonuclease

The 3’end of most m-RNAs have a polymer of Adenylate residues( 20-250) The tail prevents the attack by 3’ exonucleasesHistones and interferons do not contain poly A tails On both 5’ and 3’ end there are non coding sequences which are not translated (NCS)The intervening region between non coding sequences present between 5’ and 3’ end is called coding region. This region encodes for the synthesis of a protein.



5’ cap and 3’ tail impart stability to m RNA by protecting from specific exo nucleases.

The m- RNA molecules are formed with the help of DNA template during the process of transcription.The sequence of nucleotides in m RNA is complementary to the sequence of nucleotides on template DNA.The sequence carried on m -RNA is read in the form of codons.A codon is made up of 3 nucleotidesThe m-RNA is formed after processing of heterogeneous nuclear RNA


Heterogeneous nuclear RNA(hnRNA)

In mammalian nuclei , hnRNA is the immediate product of gene transcription The nuclear product is heterogeneous in size (Variable) and is very large.75 % of hnRNA is degraded in the nucleus, only 25% is processed to mature m RNA

Mature m –RNA is formed from primary transcript by capping, tailing, splicing and base modification.

Heterogeneous nuclear RNA(hnRNA)

Transfer RNA (t- RNA)

Transfer RNA (t- RNA)

Transfer RNA are the smallest of three major species of RNA moleculesThey have 74-95 nucleotide residuesThey transfer the amino acids from cytoplasm to the protein synthesizing machinery, hence the name t RNA.They are also called Adapter molecules, since they act as adapters for the translation of the sequence of nucleotides of the m RNA in to specific amino acidsThere are at least 20 species of t RNA one corresponding to each of the 20 amino acids required for protein synthesis.

Structural characteristics of t- RNA

1) Primary structure- The nucleotide sequence of all the t RNA molecules allows extensive intrastand complimentarity that generates a secondary structure.2) Secondary structure- Each single t- RNA shows extensive internal base pairing and acquires a clover leaf like structure. The structure is stabilized by hydrogen bonding between the bases and is a consistent feature.

Secondary structure (Clover leaf structure)

All t-RNA contain 5 main arms or loops which are as follows-a) Acceptor armb) Anticodon armc) D armd) TΨ C arme) Extra arm

Secondary structure of t- RNA Acceptor arm The acceptor arm is at 3’ end It has 7 base pairs The end sequence is unpaired Cytosine, Cytosine-

Adenine at the 3’ end The 3’ OH group terminal of Adenine binds with

carboxyl group of amino acids The t RNA bound with amino acid is called Amino

acyl t RNA CCA attachment is done post transcriptionally

b) Anticodon arm Lies at the opposite end of acceptor arm5 base pairs longRecognizes the triplet codon present in the m RNABase sequence of anticodon arm is complementary to the base sequence of m RNA codon. Due to complimentarity it can bind specifically with m RNA by hydrogen bonds.

Secondary structure of t- RNA

c) DHU arm It has 3-4 base pairsServes as the recognition site for the enzyme (amino acyl t RNA synthetase) that adds the amino acid to the acceptor arm.d) TΨC armThis arm is opposite to DHU armSince it contains pseudo uridine that is why it is so named It is involved in the binding of t RNA to the ribosomes


e) Extra arm or Variable arm About 75 % of t RNA molecules possess a short extra armIf about 3-5 base pairs are present the t-RNA is said to be belonging to class 1. Majority t -RNA belong to class 1. The t –RNA belonging to class 2 have long extra arm, 13-21 base pairs in length.


Tertiary structure of t- RNA The L shaped tertiary structure is formed by further folding of the clover leaf due to hydrogen bonds between T and D arms. The base paired double helical stems get arranged in to two double helical columns, continuous and perpendicular to one another.

Ribosomal RNA and Ribosomes

Ribosomal RNA and Ribosomes

• Ribosomal RNA (rRNA) is a component of the ribosomes, the protein synthesis factories in the cell.

• Eukaryotic ribosome (80S) made up of two dissimilar subunits.

Large subunit (60S) made up of about 50 proteins and 3 types of rRNA (5S, 5.8 S and 28 S rRNA)

Small subunit (40S) made up of about 35 proteins and 1 type of rRNA(18 S rRNA)

AE P

Large Subunit

Small Subunit

A - Amino acyl siteP - Peptidyl siteE - Exit site

Ribosomal RNA (rRNA)

Ribosomal RNA (rRNA)The functions of the ribosomal RNA molecules in the ribosomal particle are not fully understood, but they are necessary for ribosomal assembly and seem to play key roles in the binding of mRNA to ribosomes and its translationRecent studies suggest that an rRNA component performs the peptidyl transferase activity and thus is an enzyme (a ribozyme).

Comparisons of DNA and RNAFeature DNA RNABases

Purines Adenine, Guanine Adenine, Guanine

Pyrimidines Cytosine, Thymine Cytosine, Uracil

Sugar Deoxy-D-ribose D-Ribose

Size 103 to 109 bp • tRNA: 80 to 120 nt• mRNA: 500 to 10,000 nt• rRNA: 120 to 3,000 nt

Shape base-paired double-stranded helix Single-stranded random coils (tRNA is about 75% base paired)

Associated substances histones, protamines, enzymes, spermine etc.

Ribosomal proteins with rRNA

Cellular distribution Mostly in nucleus; some in mitochondria

Highest in cytoplasm but much in nucleus and nucleolus

Differences between RNA and DNAS.No. RNA DNA

1) Single stranded mainly except when self complementary sequences are there it

forms a double stranded structure (Hair pin structure)

Double stranded (Except for certain viral DNA s which are single stranded)

2) Ribose is the main sugar The sugar moiety is deoxy ribose

3) Pyrimidine components differ. Thymine is never found(Except

tRNA)

Thymine is always there but uracil is never found

4) Being single stranded structure- It does not follow Chargaff’s rule

It does follow Chargaff's rule. The total purine content in a double stranded DNA is always equal to

pyrimidine content.


5) RNA can be easily destroyed by alkalies to cyclic diesters of mono nucleotides.

DNA resists alkali action due to the absence of OH group at 2’ position

6) RNA is a relatively a labile molecule, undergoes easy and spontaneous

degradation

DNA is a stable molecule. The spontaneous degradation is very 2

slow. The genetic information can be stored for years together without

any change.

7) Mainly cytoplasmic, but also present in nucleus (primary transcript and small

nuclear RNA)

Mainly found in nucleus, extra nuclear DNA is found in

mitochondria, and plasmids etc

8) The base content varies from 100- 5000. The size is variable.

Millions of base pairs are there depending upon the organism


9) There are various types of RNA – mRNA, r RNA, t RNA, Sn RNA, Si RNA,

mi RNA and hn RNA. These RNAs perform different and specific

functions.

DNA is always of one type and performs the function of storage and

transfer of genetic information.

10) RNA is synthesized from DNA, it can not form DNA(except by the action of reverse transcriptase). It can not

duplicate (except in certain viruses where it is a genomic material )

DNA can form DNA by replication, it can also form RNA by transcription.

11) Many copies of RNA are present per cell

Single copy of DNA is present per cell.

Proteins/ Gene Expression

Proteins/ Gene Expression • Proteins make up all living materials

• Proteins are composed of amino acids – there are 20 different amino acids

• Different proteins are made by combining these 20 amino acids in different combinations

• Proteins are manufactured in the ribosomes under the strict control and order of DNA.

• The DNA language is made up of letters which are ATGCATATGGAATCAG

• These letters forms Words• ATC GCA GGA AUU AUG

• These words make sentences

•

Protein Synthesis

DNA and Protein Synthesis• DNA contains the genetic information to make amino acids•Amino acids combine to make proteins

• These proteins determine the physical traits of an organism and control cellular functions.

• Proteins do everything, and DNA gets all the credit!

Transcription is the Reading of the DNA and Changing the code to mRNA.

Translation is changing the mRNA into a trait by using tRNA to interpret the mRNA.

Transcription & Translation

• DNA has regions of coding and non-coding. The regions of DNA that code for proteins or traits are called EXONS, while the regions that do not code for proteins are called INTRONS.

Introns & Exons

Making a Protein—Transcription•First Step: Copying of genetic information from DNA to RNA called Transcription

•Part of DNA temporarily unzips and is used as a template to assemble complementary nucleotides into messenger RNA (mRNA).

• Transcription occurs inside the nucleus in a two step sequence of events.– Pre-mRNA includes both introns and exons for the

gene.– mRNA is only the coding portion (exons).

• Translation occurs in the cytoplasm at the ribosomes.– Reminder: There are three (3) types of RNA• Messenager (mRNA)• Transfer (tRNA)• Ribsomal (rRNA)

Site of Transcription&Translation

A U G G G C U U A A A G C A G U G C A C G U U

This is a molecule of messenger RNA.It was made in the nucleus by transcription from a

DNA molecule.

mRNA molecule

Codon

Messenger RNA


A ribosome on the rough endoplasmic reticulum attaches to the mRNA

molecule.

Ribosome

Ribosome


It brings an amino acid to the first three bases (codon) on the mRNA.

Amino acid

tRNA molecule

anticodon

U A C

A transfer RNA molecule arrives.

The three unpaired bases (anticodon) on the tRNA link up with the codon.

Transfer RNA


Another tRNA molecule comes into place, bringing a second amino acid.

U A C C C G

Its anticodon links up with the second codon on the mRNA.

Transfer RNA


A peptide bond forms between the two amino acids.

Peptide bond

C C G U A C

Transfer RNA


The first tRNA molecule releases its amino acid and moves off into the cytoplasm.

C C G U A C

Transfer RNA

A U G G G C U U A A A G C A G U G C A C G U U C C G

The ribosome moves along the mRNA to the next codon.

Transfer RNA


Another tRNA molecule brings the next amino acid into place.

C C G

A A U


A peptide bond joins the second and third amino acids to form a polypeptide chain.

C C G C C G

Polypeptide Chain


The polypeptide chain gets longer.

G U C

A C G

The process continues.

This continues until a termination (stop) codon is reached.

The polypeptide is then complete.

Termination (stop) codon

DNA and Protein Synthesis - Summary

Use the codon charts on the next page to find the amino acid sequence coded for by the following mRNA strands.

CAC/CCA/UGG/UGA

___________/___________/___________/____________

AUG/AAC/GAC/UAA

___________/___________/___________/____________

CAC/CCA/UGG/UGA

___________/___________/___________/____________Histidine Proline Tryptophan Stop

AUG/AAC/GAC/UAA

___________/___________/___________/____________Methionine Asparagine Aspartic Acid Stop

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