The Blueprint of Life, From DNA to Protein Chapter 7

The Blueprint of Life Characteristics of each cell dictated by information contained on DNA DNA holds master blueprint All cell structures and processes directed by DNA

Overview Complete set of genetic information referred to as genome Genome of all cells is composed of DNA Some viruses have RNA genome Functional unit of genome is the gene Gene codes for gene product Gene product is most commonly protein Study of transfer of genes is genetics Study of sequence of DNA is genomic

Overview Living cells must accomplish two general tasks to multiply DNA replication DNA expression (gene expression) Expression involves two process Transcription Copies information in DNA to RNA Translation Interpret RNA to synthesize protein Flow of information from DNA to RNA to protein Central dogma of molecular biology

Overview Characteristics of DNA Made up of deoxy-ribonucleotides Nucleotides include: Phosphate group 5 carbon sugar Deoxyribose Nucleotides bond covalently between the 5’PO4 of one nucleotide and the 3’OH of another Joining of nucleotides creates an alternating sugar-phosphate backbone

Overview Characteristics of DNA Each sugar (deoxyribose) molecule is connected to a nitrogenous base Nitrogenous bases Adenine (A) - purine Thymine (T) - pyrimidine Guanine (G) - purine Cytosine (C) – pyrimidine

Overview Characteristics of DNA Chemical structure and joining of nucleotide subunits causes strands to differ at the ends One strand has a phosphate attached at the number 5 carbon of the sugar. Termed the five prime (5’) end The other strand has a hydroxyl group attached to the number 3 carbon of the sugar. Termed the three prime (3’) end

Overview Characteristics of DNA DNA occurs as double-stranded molecule Strands are complementary to each other Due to the specific base pairing of bases A:T C:G Strands are held together with hydrogen bonds Specific hydrogen bonding between bases A is bound to T by two hydrogen bonds G is bound to C by three hydrogen bond

Overview Characteristics of DNA DNA molecule is antiparallel Strands are oriented in opposite directions Strands differ at the ends One strand oriented in the 5’ to 3’ direction. The other strand is oriented in the 3’ to 5’ direction.

Overview Characteristics of RNA RNA is made up of nucleotides Ribonucleotides RNA contains nitrogenous bases Adenine Guanine Cytosine Uracil Uracil replaces thymine in RNA RNA usually exists as single stranded molecule

Overview Characteristics of RNA Portion of DNA acts of template for RNA synthesis RNA molecule called transcript Numerous transcripts can be produced from one chromosome Either strand of DNA can act as template Three different functional groups of RNA Messenger RNA (mRNA) Ribosomal RNA (rRNA) Transfer RNA (tRNA)

Overview Regulating the expression of genes Nucleotide sequence codes for regulation mechanism for gene expression Mechanisms determine duration of synthesis of gene products Products are only made when required Key mechanism is regulation of mRNA synthesis from DNA Regulation of transcription

DNA Replication DNA is replicated to create second copy of molecule Molecule is identical to original Replication is bidirectional Replication begins at specific starting point Proceeds in opposite directions Allows replication to proceed more quickly Bi-directional replication

DNA Replication DNA replication The two strands are unwound and separated Free, unbound nucleotides match up to the newly separated nitrogenous bases of the parent strand The parent strand is also called the template strand

DNA Replication DNA replication Base pairing is specific in DNA replication Where adenine is present only thymine binds in the new strand and vice versa Where guanine is present only cytosine binds in the new strand and vice versa Bases that are improperly inserted are removed and replaced with the correct base Newly added bases are added by the enzyme DNA polymerase

DNA Replication Specifics of DNA replication As the strands of DNA unwind, it creates an area of replication called the replication fork As nucleotides are added, the replication fork moves down the parental strand

DNA Replication Specifics of DNA replication DNA polymerase adds new nucleotides as they become available. DNA polymerase can only add nucleotides to the free hydroxyl at the 3’ end DNA polymerase replicates in 5’ to 3’ direction Enzymes READS DNA template in 3’ to 5’ direction Because of the antiparallel nature of the strands of DNA, the two new strands will grow in opposite directions One strand is the leading strand One strand is the lagging strand

DNA Replication Specifics of DNA replication Leading strand Is synthesized CONTINUOUSLY as the DNA polymerase moves towards the replication fork Lagging strand Is synthesized DISCONTINUOUSLY in pieces as DNA polymerase moves away from the replication fork

DNA Replication Specifics of DNA replication DNA polymerase must bind to an RNA primer to begin synthesis A second DNA polymerase removes any RNA primers An RNA primer is required at each newly synthesized section of the lagging strand DNA ligase joins the fragments of the lagging strand

DNA Replication Specifics of DNA replication Replication is completed when the replication fork reaches the end of the parent strands The original parent strand and the newly synthesized daughter strand rewind Each new strand of DNA consists of one parent strand and one daughter strand DNA replication is referred to as semiconservative DNA Replication

Gene Expression Involves two separate but interrelated process Transcription Process of synthesizing RNA from DNA template Translation RNA is deciphered to synthesize protein

Gene Expression Transcription Transcription is the synthesis of a strand of mRNA from a DNA template mRNA carries the coded information from DNA to the ribosome, which is the site of protein synthesis mRNA also plays an important role in translation

Gene Expression Transcription During transcription the enzyme, RNA polymerase, synthesizes a complementary strand of mRNA from a portion of unwound DNA

Gene Expression Specifics of Transcription RNA polymerase binds to a region of the DNA called the promoter Only one strand of DNA acts as a template This is called the sense strand The strand not transcribed is the nonsense strand

Genet Expression Specifics of transcription Nucleotides in RNA are the same as those in DNA with one exception Thymine is replaced with uracil Binding in RNA is A:U or U:A C:G or G:C

Gene Expression Specifics of transcription RNA polymerase continues down strand of DNA until it reaches a site on DNA called the terminator At the terminator RNA polymerase and the new strand of mRNA are released from strand of DNA Transcription

Gene Expression Translation Translation is the decoding of information held in the mRNA to synthesize proteins Two more RNA molecules become involved in translation Ribosomal RNA (rRNA) Transfer RNA (tRNA)

Gene Expression rRNA forms part of the ribosomal machinery used in protein synthesis rRNA builds the ribosomes tRNA recognizes specific sequences of mRNA and transports the required amino acids to form a polypeptide chain

Gene Expression Translation The language of mRNA is in the form of codons Codons are groups of three nucleotides situated next to each other on DNA Codons are written in terms of their base sequence in mRNA The sequence of codons determines the sequence of amino acids in the protein

Gene Expression Translation There are 64 codons that make up the “alphabet” of proteins Of the 64 codons, 61 are sense codons Each coding a specific amino acid The remaining 3 are nonsense codons These code for termination of the message Codons contained in mRNA are read into proteins through translation The site of translation is the ribosome

Gene Expression In response to each codon, tRNA brings the appropriate amino acid to the site of translation Each codon has an anticodon The anticodon is complementary sequence to the codon

Gene Expression Translation Ribosomes The 30s and the 50s ribosomal subunits join together around the mRNA The ribosomes direct the binding of tRNA to the correct codon on the mRNA tRNA binds to the P site and the A site of the 50s ribosomal subunit The ribosomes bind to the mRNA to be translated

Gene Expression Specifics of Translation P site A site Specifics of Translation The first tRNA binds to a start codon in the P site of the ribosome AUG is the start codon for EVERY protein AUG codes for the amino acid methionine When the second tRNA binds to the A site, the amino acid of the first tRNA forms a peptide bond with the amino acid of the second tRNA

Gene Expression Specifics of translation After the peptide bond is formed between the two amino acids, the tRNA P site leaves the ribosome The ribosome moves distance of one codon Amino acid in the A site moves to the P site A new tRNA fills the now empty A site

Gene Expression Specifics of translation The ribosome continues down the strand of mRNA Amino acids form peptide bonds along the way Translation is terminated when the ribosomes come to a stop or nonsense codon At this point the ribosomes separate The new polypeptide chain is released The ribosome and the mRNA are free to begin translation again

Gene Expression Specifics of translation As the ribosome moves down the strand of mRNA, the start codon is exposed Once exposed, a new ribosome will attach and begin another polypeptide chain Translation

Regulation of Gene Expression Microorganisms posses mechanism to synthesize maximum amount of cell material from limited energy Controls directed at metabolic pathways Two general mechanism Allosteric inhibition of enzymes Controlling synthesis of enzymes Directed at making only what is required

Regulation of Gene Expression Principles of regulation Not all genes subjected to regulation Enzymes can be classified according to characteristics of regulation Constitutive enzymes Constantly synthesized Enzymes of glycolysis Inducible enzymes Not regularly produced turned on in certain conditions Β-galactosidase Repressible enzymes Routinely synthesized Generally involved in biosynthesis

Regulation of Gene Expression Mechanisms controlling transcription Often controlled by regulatory region near promoter Protein binds to region and acts as “on/off” switch Binding protein can act as repressor or activator Repressor blocks transcription Activator facilitates transcription Set of genes controlled by protein is called an operon

Regulation of Gene Expression Repressors Control mechanism that inhibits gene expression and decreases the synthesis of enzymes Repression is usually in response to the overabundance of an end product Repression decreases the rate synthesis of enzymes leading to the formation of the particular end product Regulatory proteins called repressors mediate repression Repressors block the ability of RNA polymerase to bind and initiate protein synthesis

Regulation of Gene Expression Activators Control mechanism that turns on the transcription of a gene or set of genes Inducers are substances that act to induce transcription Enzymes synthesized in the presence of inducers are called inducible enzymes

Regulation of Gene Expression Operon model of gene expression An operon is a set of genes that includes an operator, promoter and structural genes An operon is divided into two regions, the control region and the structural region The control region include the operator and the promoter This region controls transcription The operator acts as the “on-off” switch The structural region includes the structural genes This region contains the genes being transcribed

Operon structure Promoter – Binding site for RNA polymerase Operator Gene 1 Gene 3 Gene 2 Promoter Promoter – Binding site for RNA polymerase Operator – binding site for the repressor protein for the regulation of gene expression Structural Genes – DNA sequence for specific proteins

Regulation of Gene Expression Lac operon Example of induction of gene expression Near the operon on the DNA is a regulatory gene called the “I” gene This codes for the repressor protein When lactose is absent, the repressor protein binds to the operator gene Binding of the repressor gene prevents RNA polymerase from transcribing the structural genes No mRNA is made and no enzymes are synthesized

Regulation of Gene Expression Lac operon When lactose is present the repressor binds to lactose instead of the operator With the repressor bound to lactose, RNA polymerase is able to bind to the promoter and transcribes the structural genes Lactose acts as an inducer by keeping the repressor from binding to the operator It induces the transcription of the structural genes

Lac Operon 1. 2. 3. Lac Operon Operator Gene 1 Gene 3 Gene 2 Promoter RNA polymerase Repressor 2. 3. Lac Operon Lactose

Gene Expression and Environmental Fluctuations Many organisms adapt to changing environments by altering level of gene expression Mechanisms include Signal transduction Natural selection

Gene Expression and Environmental Fluctuations Signal transduction Process that transmits information from external environment to inside cell Allows cell to respond to changes Two-component regulatory systems Relies on sensor and response regulator proteins Sensors recognize change in environment Response regulators activate or repress gene expression Quorum sensing Organisms sense density of population Enables activation of genes beneficial to the mass

Gene Expression and Environmental Fluctuations Natural selection Mechanisms to enhance survivability Antigenic variation Alteration in characteristics of certain surface proteins Example: Neisseria gonorrhoeae hides from host immunity by changing numerous surface proteins Phase variation Routine switching on and off of certain genes Altering expression allows portions of population to survive and multiply