chapter 8 2770 Key concepts

Genes control phenotypic traits through

the expression of proteins

The DNA of a gene is

a linear sequence of many nucleotides

The DNA of a gene is a linear sequence of many nucleotides

This is transcribed into RNA

• Which is translated into a polypeptide

a gene is defined as

a region of DNA that can be expressed

Transcription

The synthesis of RNA under the direction of DNA

Translation

The synthesis of proteins under the direction of RNA

Central Dogma of Molecular Biology

DNA → RNA → Polypeptides (Protein)

Transcription Translation

DNA

Long term information storage and the molecule of heredity

RNA

• Short term information transmission and processing

• mRNA is the intermediate if the product is a protein/polypeptide

Protein

• Typically, the end product, the functional molecule

Genetics

• The study of genes

• How they carry information, how information is expressed, and how genes are replicated

Chromosomes

• Structures containing DNA that physically carry hereditary information

• Chromosomes contain genes

Genes

• Segments of DNA that encode functional products, usually proteins

Genome

• All the genetic information in a cell

Genomics

• Sequencing and molecular characterization of genomes

Allele

• Alternative versions of a gene

Genotype

• The allelic makeup of an organism (hetero/homozygous)

• Phenotype

• The outward representation of the genotype (morphology)

• Largely based on the proteins being expressed

• Changes in genotype can lead to changes in phenotype

Bacteria usually have a

single circular chromosome

Genome consists of

• Protein-encoding Genes

• Noncoding regions

The Flow of Genetic Information

• Expression

• Vertical Gene Transfer

• Horizontal Gene Transfer

DNA forms a

double helix

double helix

"Backbone" consists of deoxyribose-phosphate

• Two strands of nucleotides are held together by H-bonds

• Between A-T and C-G

• Strands are antiparallel (oriented in opposite directions)

Bacteria pack their DNA into a series of

loops or domains

Positive supercoils -

DNA is overwound

• Negative supercoils -

DNA is underwound Relative to the relaxed number of helical turns in the strand

Most cells possess

negatively supercoiled DNA

DNA strands are easier to separate when

negatively supercoiled

Archaea living in hot, acidic environments

positively supercoil DNA

Topoisomerases

Enzymes that maintain the proper level of supercoiling

Topoisomerase I

Reduce the number of negative supercoils

Topoisomerase II

Introduce negative supercoils into DNA

DNA Gyrase and Type II topoisomerase

common to bacteria

• Though it is found in some protists (plastids)

Target for several antibiotic drugs

Topoisomerase and gyrase

relax the strands

Helicase

separates the strands

DNA Helicase

• Separates strands of the double helix

• Expanding the replication bubble

Primase

• Synthesizes a short complementary RNA

• Based on the ssDNA template

• DNA pol III

Allows second DNA pol to attach

Nucleoside triphosphates are added

• DNA nucleotides with 3 phosphates

• ⍺ phosphate forms a bond with the 3'-OH

• Forming the phosphodiester bond (C-O-P-O-C)

• Hydrolysis cleaves the two distal phosphates

• Releases as pyrophosphate (P2O7) [energetically drives the reaction]

• Hydrolysis of pyrophosphate effectively locks the bond in place

• DNA pol III cannot polymerize without an -OH target

• Which is why primers are required to begin synthesis

DNA pol III cannot polymerize without an -OH target

Which is why primers are required to begin synthesis

DNA polymerases rely on

complementary base pairing

DNA polymerases rely on complementary base pairing

This ensures that the nucleotide being added is appropriate

Issues with complementary pairing typically cause rejection of the nucleotide

The polymerase then brings in another that fits properly

Ribonucleic acid

• Contains ribose

• Is single-stranded

Adenine hydrogen bonds

Uracil

• Cytosine hydrogen bonds

Guanine

Messenger RNA (mRNA)

Encodes the amino acid sequence of one or more proteins

Ribosomal RNA (rRNA)

Catalytic component of ribosomes, catalyzes peptide bonds

Transfer RNA (tRNA)

Carry amino acids and an anticodon which is used to match with an mRNA codon

Transcription

Conducted by multi-subunit DNA-dependent RNA polymerases

Genes are marked by sequences which allow their recognition

Allows the cell to only express gene regions, and allow for regulation

Promotor

Sequence upstream of a gene, attracts sigma factor to begin transcription

Initiation

• RNA pol holoenzyme binds to the promoter

• This is followed by melting of the helix

Elongation

• The RNA chain is extended one base at a time

Termination

• RNA pol detaches from the DNA, after the transcript is made

RNA polymerase holoenzyme binds DNA

• Scans along DNA looking for a promotor sequence • Forms a loosely bonded closed-complex with DNA

• The DNA strands are separated to form an open complex

• Through the unwinding of one helical turn

• In the open-complex the DNA is tightly bound

• Which causes transcription to begin

Transcription stops when

it reaches the terminator sequence on DNA

Terminators are often sequences

which are difficult to copy

Transcription produces

an mRNA copy of the gene

• Now that sequence must be used to produce a protein

Genetic information is written in a genetic code

Which consists of 3 nucleotide long "codons" (3 x 4 bases = 64 codons)

• Each codon codes for one of the 20 amino acid

Base Triplet

• A sequence of three DNA nucleotides

• Each triplet represents one amino acid

Codon

The 3-base sequence in mRNA

64 possible codons available to represent

the 20 amino acids

Wobble Position

Less stringent pairing between bases of codon and the tRNA anticodon

Open Reading Frame (ORF)

The region between translation start codon and stop codon, gene coding region

Ribosome

• Molecular machine responsible for protein formation

• Translates the language of the mRNA code into the amino acid sequences

• Catalyzes the formation of peptide bonds

• Composed of two subunits, each of which includes rRNA and proteins

The ribosome has

multiple tRNA-binding sites

A (Aminoacyl) Site

• Hosts the tRNA carrying the incoming amino acid

P (Peptidyl) Site

• Hosts the tRNA attached to the growing peptide chain

E (Exit) Site

• Hosts the tRNA that carried the previous amino acid added

• The uncharged tRNA which must be released and recharged

The correct reading frame is marked on the

mRNA

If an mRNA contains multiple genes, there will be multiple

RBSs

Initiation

• Brings the two ribosomal subunits together with the mRNA and first tRNA

• Placing the first amino acid in position

Elongation

• Sequentially adds amino acids as directed by mRNA transcript

Termination

• Releases the completed protein and recycles ribosomal subunits

In bacteria, translation can begin before transcription is complete

• This happens because transcription occurs in the cytoplasm

• Polyribosome

• A group of ribosomes bound to and translating a single mRNA

In eukaryotes, transcription occurs in the

nucleus

While translation occurs in the

cytoplasm

Post-Transcriptional Modifications

Most eukaryotic mRNAs also contain noncoding intron sequences

• Sequences in their DNA which are not used in the production of protein

• These are not found in bacteria, but are found in some archaeans

Introns (Intragenic Regions)

Do not code for amino acids (non-coding)

Exons (Expressed Regions)

• Code for amino acid sequence

Splicing

Introns must be removed to form a functional protein

Small Nuclear Ribonucleoproteins (snRNPs)

• Remove introns and splice exons together

Metabolic Regulation

Regulation is important for cellular health

Regulation is important for cellular health

• Prevents wasteful production of products that are unneeded

• The cell has only so much space, energy and nutrients

Metabolic Regulation

• Allows the cell to activate pathways when they are needed

• Allows the cell to activate pathways when they are needed

• Activate viral defenses when the cell is under attack

• Activate cell division proteins when it is time to divide

Some products could interfere with operations of others

• Activating cell division mechanisms and sporulation mechanisms would be disastrous

Constitutive Genes

Are expressed at a fixed rate

• These do not appear to be regulated and are always "turned on

• Inducible genes

• Induction turns on expression, default position of an inducible gene is off • Initiated by an inducer

Repressible genes

• Repression inhibits expression, default position of a repressible gene is on

• Mediated by repressors, proteins that block transcription

• Catabolite repression

• Inhibits cells from using carbon sources other than glucose

Promoter

• Segment of DNA where RNA polymerase initiates transcription of genes

Operator

• Segment of DNA that controls transcription of structural genes

Operon

Set of operator and promoter sites and the structural genes they control

In an inducible operon

• Structural genes are not transcribed unless an inducer is present

• Proteins encoded by the lac operon are needed to

metabolize lactose

Lac I: inhibitor proteins specific to lacZYA

• Encoded by its own gene and operon

With no lactose, lac I represses

lacZYA

• Binds to the operator blocking translation