Central dogma I and IICentral dogma I and IIthe flow of genetic information

1 Th T f i1. The Transforming Principle

2. Overview of Central DogmaDogma

3. Nucleic Acid Structure4. The Organization of

DNA i C llDNA in Cells5. DNA Replication6. Gene Structure and the

Genetic Code7. Transcription8. Translation9. Post-Translational

Modification

DNA as Genetic MaterialTransformation – Transforming principle

G iffith i 1928 b d th h f i l t• Griffith in 1928 observed the change of non-virulent organisms into virulent ones as a result of “transformation”– MacLeod and McCarty in 1944 showed that the

transforming principle was DNAtransforming principle was DNA

Transforming principleTransforming principle

The Flow of Genetic InformationThe Flow of Genetic Information

DNA stores genetic– DNA stores genetic information

– Information is d li t d b li ti

DNA GenotypeReplication

duplicated by replication and is passed on to next generation

RNA

Transcription

– transcription yields a ribonucleic acid (RNA) copy of specific genes

Polypeptide

Translation

– translation uses information in messenger RNA (mRNA) to synthesize a

l tid

yp p

Protein

Posttranslational modification

polypeptide• Also involves

activities of transfer RNA (tRNA) and ib l RNA Cell

Phenotype

ribosomal RNA (rRNA)

Cell

Nucleic Acid StructureTh l i id DNA d RNA l f l tid• The nucleic acids, DNA and RNA are polymers of nucleotides– linked together by phosphodiester bonds

• DNA and RNA differ inDNA and RNA differ in– the nitrogenous bases they contain– the sugars they contain– whether they are single or double stranded

Deoxyribonucleic Acid (DNA)y ( )• Polymer of nucleotides• Contains the bases adenine, guanine, cytosine and thymine

S• Sugar is deoxyribose• Molecule is usually double stranded• Base pairing

– Adenine (purine) and thymine (pyrimidine) pair by 2Adenine (purine) and thymine (pyrimidine) pair by 2 hydrogen bonds

– Guanine (purine) and cytosine (pyrimidine) pair by 3 hydrogen bonds

Ribonucleic Acid (RNA)Ribonucleic Acid (RNA)

P l f l tid Th diff t t• Polymer of nucleotides

• Contains the bases

• Three different types which differ from each other in function, site of synthesis (in eucaryoticadenine, guanine,

cytosine and uracilsynthesis (in eucaryotic cells) and in structure– messenger RNA

(mRNA)• Sugar is ribose

• Most RNA molecules

(mRNA)– ribosomal RNA

(rRNA)os o ecu esare single stranded – transfer RNA

(tRNA)

The Organization of DNA in CellsThe Organization of DNA in Cells

I ll A h d• In all Archaea and most bacteria DNA is a circular double helix

• Further twisting results inresults in supercoiled DNA– In bacteria the

DNA is associatedDNA is associated with basic proteins

• Help organize the DNA into a coiledDNA into a coiled chromatin like structure

DNA FormsDNA Forms

Eucaryotic DNA OrganizationEucaryotic DNA Organization

• DNA is more• DNA is more highly organized in eucaryotic chromatin where it is associated with histones,with histones, small basic proteins

• The combination of DNA and proteins isproteins is called a nucleosome

DNA ReplicationDNA Replication• Complex process• Complex process

involving numerous proteins which help ensure accuracy

• The 2 strands separate, each serving as a t l t f th itemplate for synthesis of a complementary strand

• Synthesis is semi-conservative; each daughter cell obtains

ld done old and one new strand

In most procaryotes bidirectional from a single origin of replication

Rolling Circle ReplicationRolling Circle Replication

ll• some small circular genomes (e.g., viruses and (e.g., viruses and plasmids– replicated by

lli i lrolling-circle replication

Eucaryotic DNA ReplicationEucaryotic DNA Replication• eucaryotic DNA is ~1,400 times longer than

ti DNA d i liprocaryotic DNA and is linear

• many replication forks are used simultaneously with y p ymany replicons present

1. Ori2 Helicase

6. RNA primer7 DNA polymerase2. Helicase

3. DNA Gyrase4 SSB

7. DNA polymerase8. Leading/lagging strand9 DNA ligase4. SSB

5. Primase9. DNA ligase10. Termination

DNA PolymeraseDNA Polymerase

Proofreading, removal of mismatched base from 3’ end of growing strand by exonuclease activity of enzyme

• Gene, defined as the nucleic acid sequence that codes for a polypeptide, tRNA or rRNA

• Template strand directs RNA synthesis (3’ to 5’ direction)Template strand directs RNA synthesis (3 to 5 direction)• Promoter is located at the start of the gene and the binding

site for RNA polymerase• Leader sequence is transcribed into mRNA but is not

translated into amino acidstranslated into amino acids• Shine-Delgarno sequence important for initiation of translation• reading frame, organization of codons such that they can be

read to give rise to a gene product

Genetic CodeGenetic Code• code degeneracy

– up to six different codons can code for a single amino acidd• sense codons

– the 61 codons that specify amino acids• stop (nonsense) codons

the three codons used as translation termination signals– the three codons used as translation termination signals– do not encode amino acids

Importance of reading frame

Transcription• RNA synthesis under the direction of DNA

– RNA produced has complementary sequence to the template DNAf– Three types of RNA are produced

• mRNA carries the message for protein synthesis• tRNA carries amino acids during protein synthesis• rRNA molecules are components of ribosomes

• Polygenic mRNA often found in bacteria and archaea– contains directions for > 1 polypeptide

• Catalyzed by a single RNA polymerase– Reaction similar to that catalyzed by DNA polymerase

• ATP,GTP,CTP and UTP are used to produce a complementary RNA copy of the template DNA sequence

Transcription in procaryotesTranscription in procaryotes

• Initiation• Initiation• Elongation• Termination

- the sigma factor has no catalytic activityno catalytic activity but helps the core enzyme recognize the start of genes

– holoenzyme = core enzyme + sigma factorfactor

• Only the holoenzyme can begin transcription

Promoter- site where RNA polymerase binds to initiatesite where RNA polymerase binds to initiate

transcription

The Transcription Bubble (elongation)

• After binding, RNA polymerase unwinds the DNA

• Transcription bubble produced– Moves with the

polymerase as it p ytranscribes mRNA from template strand

• Within the bubble a temporary RNA:DNA hybrid is formed

Transcription TerminationTranscription TerminationOcc rs hen• Occurs when core RNA polymerase dissociates from template DNA

• DNA sequences qmark the end of gene in the trailer and the terminatorterminator

• Some terminatorsterminators require the aid of the rho factor for termination

Transcription in EucaryotesTranscription in Eucaryotes• Several RNA polymerases

P t diff f• eucaryotic genes

h ( d– Promotes differ from those in bacteria by having combinations of many elementsCatalyzes production of

– have exons (expressed sequences) and introns (intervening sequences) that code for RNA that is never translated into– Catalyzes production of

heterogeneous nuclear RNA (hnRNA) which undergoes posttranscriptional

is never translated into protein

posttranscriptional modification to generate mRNA

Transcription in the ArchaeaTranscription in the Archaea• RNA polymerase has similarities to both p y

bacteria and eucaryotic enzyme

• similarities with eucaryotes– archaeal gene promoters and binding of the

RNA polymeraseRNA polymerase – introns present in some archaeal genes

• similarities with procaryotes– mRNA is polygenicmRNA is polygenic

TranslationTranslation• synthesis of polypeptide directed by sequence of nucleotides

in mRNA• direction of synthesis N terminal C-terminal• ribosome

– site of translation– polyribosome – complex of mRNA with several ribosomes

The Ribosome• Procaryotes, 70S ribosomes = 30S + 50S subunits• Eucaryotes, 80S ribosomes = 40S + 60S subunits• mitochondrial and chloroplast ribosomes resemble procaryotic• mitochondrial and chloroplast ribosomes resemble procaryotic

ribosomes• peptidyl (donor; P) site, binds initiator tRNA or tRNA attached

to growing polypeptide (peptidyl-tRNA)• aminoacyl (acceptor; A) site, binds incoming aminoacyl-tRNA• exit (E) site, briefly binds empty tRNA before it leaves

ribosome

Aminoacyl tRNAAminoacyl-tRNA

tt h t f• attachment of amino acid to tRNA

• catalyzed by aminoacyl-tRNA

th tsynthetases– at least 20

• each specific for psingle amino acid and for all the tRNAs to which each may beeach may be properly attached (cognate tRNAs)

Coupled Transcription and Translation in Procaryotes