Prokaryote Cell Prokaryote Cell Structure and FunctionStructure and Function

Background and Background and ClassificationClassification

CaulobacterCaulobacter

crescentuscrescentus

Prokaryote CellsProkaryote Cells

No nuclear membraneNo nuclear membraneNo cellular organelles( membrane No cellular organelles( membrane

bound organelles)bound organelles)Ribosomal sizeRibosomal sizeDNADNARNARNASizeSizeCell wall and cell membrane Cell wall and cell membrane

A New View of LifeA New View of Life Three Domains of lifeThree Domains of life Carl Woese Carl Woese

responsible for responsible for elucidating specific elucidating specific DNA differences DNA differences between the between the prokaryotesprokaryotes

Looked at the Looked at the relationship between relationship between the organisms and the organisms and created a branching created a branching tree( see chart)tree( see chart)

Carl WoeseCarl Woese Studied the molecular biology of the Studied the molecular biology of the

prokaryotesprokaryotes Used 16s rRNA’s to create his Tree of Life - Used 16s rRNA’s to create his Tree of Life -

this is interpreted as an evolutionary this is interpreted as an evolutionary distance between types of bacteria in distance between types of bacteria in terms of differences in the 16s rRNAterms of differences in the 16s rRNA

Changes in 16s rRNA may be used as a Changes in 16s rRNA may be used as a molecular chronometer or watch to convey molecular chronometer or watch to convey the time required to make changes in the the time required to make changes in the genes and proteins – ( Pauling 1965)genes and proteins – ( Pauling 1965)

Parameters used in Parameters used in classificationclassification

DNA hybridization – homology of DNA DNA hybridization – homology of DNA sequences – the use of probes( DNA sequences – the use of probes( DNA and m RNA)and m RNA)

G+C content – DNA melting curves. G+C content – DNA melting curves. DNA sequencingDNA sequencingProtein homologyProtein homologyBiochemical characteristicsBiochemical characteristicsMolecular characteristics ( expression)Molecular characteristics ( expression)

Prokaryote DomainsProkaryote DomainsSimilaritiesSimilarities Bacteria and Archaea Bacteria and Archaea have smaller ribosomeshave smaller ribosomes ( 70s)( 70s) No membrane bound No membrane bound

nucleusnucleus Generally one ds circularGenerally one ds circularchromosome- genomic DNAchromosome- genomic DNA( there are many exceptions)( there are many exceptions) Many have plasmidsMany have plasmids Operon organization and Operon organization and

gene regulation gene regulation mechanismsmechanisms

DifferencesDifferences Cell wall differences Cell wall differences

between Archaea and between Archaea and Eukarya – PeptidoglycanEukarya – Peptidoglycan

Cell membane – ester Cell membane – ester linkage versus ether linkage versus ether linkagelinkage

Ribosome sensitivity to Ribosome sensitivity to antibioticsantibiotics

( chloramphenicol and ( chloramphenicol and streptomycinstreptomycin

Ribosomal sensitivity to Ribosomal sensitivity to diptheria toxindiptheria toxin

RNA sequencesRNA sequences RNA PolymerasesRNA Polymerases

ArchaeaArchaea

Includes organisms regarded a Includes organisms regarded a extremophilesextremophiles

Methanogens Methanogens HalophilesHalophilesHyperthermophilesHyperthermophilesNitrogen bacteriaNitrogen bacteria

Classification - BacteriaClassification - Bacteria Proteobacteria – Five Classes – largest group. Proteobacteria – Five Classes – largest group.

Very diverseVery diverse Class I – Alpha proteobacteria – range from Class I – Alpha proteobacteria – range from

Nitrogen fixing bacteria vital to recycling of Nitrogen fixing bacteria vital to recycling of Nitrogen to pathogens like Nitrogen to pathogens like RickettsiaeRickettsiae

Class II – ( Betaproteobacteria) includes Class II – ( Betaproteobacteria) includes NeisseriaNeisseria species ( species ( gonnorheae and meningitidis )gonnorheae and meningitidis )

Class III( Gammaproteobacteria) includes – Class III( Gammaproteobacteria) includes – E. coli, E. coli, Salmonella, Shigella,Salmonella, Shigella, and other pathogens and other pathogens

Class IV – Organisms that are unique – Class IV – Organisms that are unique – Bdellovibrio that devours gram negative bacteriaBdellovibrio that devours gram negative bacteria

Class V – Includes Class V – Includes Campylobacter and Campylobacter and Helicobacter pyloriHelicobacter pylori

Gram Positive Bacteria – Gram Positive Bacteria – High G +C contentHigh G +C content

ActinomycesActinomyces – Bacteria that are – Bacteria that are found in the environmentfound in the environment

Mycobacterium, actinomyces, and Mycobacterium, actinomyces, and streptomycesstreptomyces

StreptomycesStreptomyces and and actinomycesactinomyces are are soil bacteria with unusual soil bacteria with unusual characteristics that have contributed characteristics that have contributed to antibiotic therapy ( Selman to antibiotic therapy ( Selman Waksman – Rutgers)Waksman – Rutgers)

SpirochetesSpirochetes

Unique organisms – Unique organisms – Treponema Treponema

pallidumpallidum BorreliaBorrelia LeptospiraLeptospira

Gram Positive Bacteria – Low Gram Positive Bacteria – Low G-C contentG-C content

Gram positive organismsGram positive organismsMedically importantMedically importantClostridiumClostridiumMycoplasmaMycoplasmaBacilli, Enterococcus, and Bacilli, Enterococcus, and

StreptococcusStreptococcus

Prokaryote – Cell SizeProkaryote – Cell Size Mycoplasmas are Mycoplasmas are

about the size of a about the size of a virus with the virus with the diameter of 0.3 diameter of 0.3 µm

E. coli is a more typical bacterium with dimensions of 1.1-1.5 µm wide by 2.0-6.0 µm in length.

The size of bacteria The size of bacteria ranges from 0.1 to ranges from 0.1 to about 600 about 600 µm over a single dimension

They are as small as the largest viruses to large enough for single cells to be visible by the naked eye

The range in sizeThe range in size

Largest greater Largest greater than 50 than 50 μm in diameter

Smallest less than .3 μm

From ultra to nanoFrom ultra to nano

Epulopiscium fishelsoni Nanobacteria

Shapes of bacteriaShapes of bacteria

RodsCurved - spirochetes

Cocci

The Prokaryote Cell The Prokaryote Cell

Prokaryote Cell StructuresProkaryote Cell Structures

Prokaryote Cell Prokaryote Cell UltrastructureUltrastructure

Cell WallCell Wall Rigid structure that lies Rigid structure that lies

just outside the plasma just outside the plasma membranemembrane

Maintains shape, protects Maintains shape, protects the membrane, and the membrane, and regulates transportregulates transport

Basic Molecular components of Basic Molecular components of the cell wallthe cell wall

PeptidoglycanPeptidoglycan is a complex polymer is a complex polymer of sugars and amino acidsof sugars and amino acids

The peptidoglycan that is unique to The peptidoglycan that is unique to bacteria is bacteria is mureinmurein. .

The fact that murein is unique has The fact that murein is unique has made it a target of antibiotics( an made it a target of antibiotics( an entire class) that inhibits the entire class) that inhibits the synthesis of the wall. ( synthesis of the wall. ( Beta lactamsBeta lactams which includes penicillin)which includes penicillin)

The basic structureThe basic structure Glycan sugar chains linked Glycan sugar chains linked

by peptides.by peptides. N-acetyl glucosamineN-acetyl glucosamine ( NAG)( NAG) and N- acetyl-muramic and N- acetyl-muramic

acid( NAM)acid( NAM) Linked by four peptide – Linked by four peptide –

third is lysinethird is lysine Cross – Linked with Cross – Linked with

glycinesglycines This structure is similar This structure is similar

throughout the Domain throughout the Domain bacteria but has variable bacteria but has variable chemical properties in chemical properties in different speciesdifferent species

PeptidoglycanPeptidoglycan This This

structure( compared structure( compared to the chain mail of to the chain mail of medieval soldiers) medieval soldiers) covers the outer covers the outer surface of the surface of the bacterial cell. This bacterial cell. This determines the shape determines the shape of the bacterium for of the bacterium for instance coccus or instance coccus or bacillusbacillus

Additional Cell Wall componentAdditional Cell Wall component

An actin like protein has been found An actin like protein has been found underlying bacterial cell walls.underlying bacterial cell walls.

Cytoskeletal elements were previously Cytoskeletal elements were previously thought to be absent from bacterial cellsthought to be absent from bacterial cells

These proteins have been found in gram These proteins have been found in gram negative bacterianegative bacteria

This new research indicates that the origin This new research indicates that the origin of the eukaryote cell cytoskeleton may be of the eukaryote cell cytoskeleton may be of prokaryote origin.of prokaryote origin.

The Two Major Types of The Two Major Types of Bacterial Cell WallsBacterial Cell Walls

Bacteria are divided into two major Bacteria are divided into two major groups based on the response to groups based on the response to Gram-stain procedure.Gram-stain procedure.gram-positive bacteria stain purplegram-positive bacteria stain purplegram-negative bacteria stain pinkgram-negative bacteria stain pink

staining reaction due to cell wall staining reaction due to cell wall structurestructure

Teichoic AcidTeichoic Acid

Teichoic acids are found in Gram Positive Cell Walls

Polymers of glycerol

or ribitol joined by phosphate groups Polymers of 30 long Extend beyond the

cell wall

Comparison of cell wall Comparison of cell wall structurestructure

The Gram Positive cell wall is characterized by a thick layer of Peptidoglycan. This causes the bacterium to stain purple with the Gram Stain

The Gram Negative cell wall has a layer of lipids overlying the Peptiodglycan layer which is much thinner.

This results in a pinkish color upon staining.

Gram Stain TechniqueGram Stain Technique

1.1. Make a smear( spread across the surface Make a smear( spread across the surface of the slideof the slide

2.2. Air dry smearAir dry smear3.3. Heat fixHeat fix4.4. Cover smear with Crystal violet – 1 Cover smear with Crystal violet – 1

minuteminute5.5. ( gram positive) – purple and rinse( gram positive) – purple and rinse6.6. Iodine( mordant) – 1 minute and rinseIodine( mordant) – 1 minute and rinse7.7. Alcohol( decolorizer) – seconds and rinseAlcohol( decolorizer) – seconds and rinse8.8. Saffranin – gram negative – pink – 1 Saffranin – gram negative – pink – 1

minute and rinseminute and rinse

Gram Staining Gram Staining

Thought to involve constriction of the Thought to involve constriction of the thick peptidoglycan layer of gram-thick peptidoglycan layer of gram-positive cellspositive cellsconstriction prevents loss of crystal constriction prevents loss of crystal

violet during decolorization stepviolet during decolorization stepThinner peptidoglycan layer of gram-Thinner peptidoglycan layer of gram-

negative bacteria does not prevent negative bacteria does not prevent loss of crystal violetloss of crystal violet

Gram PositiveGram Positive

Gram PositiveGram Positive

Gram NegativeGram Negative

Gram NegativeGram Negative

The Outer LPS - The Outer LPS - LipopolysaccharideLipopolysaccharide

consist of three consist of three partsparts lipid Alipid A core polysaccharidecore polysaccharide O side chainO side chain ( (O O

antigenantigen))

Characteristics of the Gram Characteristics of the Gram Negative Cell WallNegative Cell Wall

Protection from host defenses (O Protection from host defenses (O antigen)antigen)

Contributes to negative charge on Contributes to negative charge on cell surface (core polysaccharide)cell surface (core polysaccharide)

Helps stabilize outer membrane Helps stabilize outer membrane structure (lipid A)structure (lipid A)

LPSLPS

Lipid A is an unusual glycolipid Lipid A is an unusual glycolipid composed of a disaccharide with composed of a disaccharide with attached sort-chain fatty acids and attached sort-chain fatty acids and phosphate groups. This is linked to phosphate groups. This is linked to fever and shock invertebrates and is fever and shock invertebrates and is an endotoxinan endotoxin

LPSLPS

The core –A short series of sugars The core –A short series of sugars attached to Lipid Aattached to Lipid A

The O antigen is a long carbohydrate The O antigen is a long carbohydrate chain up to 40 sugar residues in chain up to 40 sugar residues in length which is bound to the core.length which is bound to the core.

The hydrophilic carbohydrate chains The hydrophilic carbohydrate chains of the O antigen exclude hydrophobic of the O antigen exclude hydrophobic compoundscompounds

ConnectionsConnections

Braun’s lipoproteins connect outer Braun’s lipoproteins connect outer membrane to peptidoglycanmembrane to peptidoglycan

Adhesion sitesAdhesion sitessites of direct contact (possibly true sites of direct contact (possibly true

membrane fusions) between plasma membrane fusions) between plasma membrane and outer membranemembrane and outer membrane

substances may move directly into cell substances may move directly into cell through adhesion sitesthrough adhesion sites

O antigen and importanceO antigen and importance

The O antigen is highly The O antigen is highly immunogenic. It elicits a strong immunogenic. It elicits a strong antibody response when introduced antibody response when introduced when introduced into a vertebrate when introduced into a vertebrate host.host.

E coli 157:H7 is the pathogenic form E coli 157:H7 is the pathogenic form of E. coli as compared to a of E. coli as compared to a commensal in the gut. This is commensal in the gut. This is considered to be a virulence factor.considered to be a virulence factor.

LPS - significanceLPS - significance

More permeable than plasma More permeable than plasma membrane due to presence of membrane due to presence of porin porin proteinsproteins and transporter proteins and transporter proteinsPorin proteins form channels through Porin proteins form channels through

which small molecules (600-700 daltons) which small molecules (600-700 daltons) can passcan pass

These proteins and their channels are of These proteins and their channels are of great complexitygreat complexity

Larger molecules are translocated by Larger molecules are translocated by specialized protein complexesspecialized protein complexes

Periplasmic spacePeriplasmic space The two cell wall structures create an The two cell wall structures create an

internal compartment is the periplasminternal compartment is the periplasm This compartment contains degradative This compartment contains degradative

enzymes such as nucleases, proteases, and enzymes such as nucleases, proteases, and phosphatasesphosphatases

Binding proteins that have a high affinity for Binding proteins that have a high affinity for amino acids and sugars are also presentamino acids and sugars are also present

It is space that contains the Beta lactamases It is space that contains the Beta lactamases that degrade antibiotics so that they cannot that degrade antibiotics so that they cannot interfere with the cell wall synthesisinterfere with the cell wall synthesis

Function of LPS and cell wallFunction of LPS and cell wall

Osmotic lysisOsmotic lysisCan occur when cells are in hypotonic Can occur when cells are in hypotonic

solutionssolutionsMovement of water into cell causes Movement of water into cell causes

swelling and lysis due to osmotic swelling and lysis due to osmotic pressurepressure

Cell wall protects against osmotic Cell wall protects against osmotic lysislysis

Plasmolysis and LysisPlasmolysis and Lysis

PlasmolysisPlasmolysis useful in food useful in food

preservationpreservation e.g., dried foods e.g., dried foods

and jelliesand jellies Osmotic lysisOsmotic lysis

basis of basis of lysozymelysozyme and and penicillinpenicillin action action

Osmotic lysisOsmotic lysis can occur when cells can occur when cells

are in hypotonic are in hypotonic solutionssolutions

movement of water movement of water into cell causes into cell causes swelling and lysis due swelling and lysis due to osmotic pressureto osmotic pressure

Cell wall protects Cell wall protects against osmotic lysisagainst osmotic lysis

protoplast – the absence ot cell walls in gram-positive

spheroplast – the absence of a cell wall in gram-negative

Acid Fast Cell WallAcid Fast Cell Wallhttp://student.ccbcmd.edu/courses/bio141/lecguide/unit1/proshttp://student.ccbcmd.edu/courses/bio141/lecguide/unit1/prostruct/afcw.htmltruct/afcw.html

Mycobacterium tuberculosisMycobacterium tuberculosis is a pathogen that has a is a pathogen that has a different solutiondifferent solution

Their cell walls contain waxes Their cell walls contain waxes known as mycolic acidsknown as mycolic acids

These molecules are arranged These molecules are arranged in two layers( hydrophilic tails in two layers( hydrophilic tails between them)between them)

These are attached to the These are attached to the Peptidoglycans cell wall and Peptidoglycans cell wall and form thick layers around the form thick layers around the exteriorexterior

Proteins are interspersed Proteins are interspersed within and enable nutrients to within and enable nutrients to pass throughpass through

Acid Fact BacteriaAcid Fact Bacteria

Acid fast stain – Acid fast stain – The outer covering The outer covering

is unaffected by is unaffected by hydrochloric acid hydrochloric acid which resulted in which resulted in the name, acid fastthe name, acid fast

Acid fast bacilli Acid fast bacilli stain red due to stain red due to carbol fuschincarbol fuschin

Characteristics of the acid fast Characteristics of the acid fast cell wallcell wall

Outer waxy layer resists phagocytes Outer waxy layer resists phagocytes and avoids the immune systemand avoids the immune system

The permeability to nutrients is The permeability to nutrients is minimal so that growth is very slowminimal so that growth is very slow

Mycobacterium tuberculosis may Mycobacterium tuberculosis may divide only once in 24 hoursdivide only once in 24 hours

Other variantsOther variants

Mycoplasmas Mycoplasmas are bacteria that lack are bacteria that lack cell walls cell walls

Mycoplasma pneumoniae Mycoplasma pneumoniae contain contain sterols in the membranes which sterols in the membranes which protects against swelling and lysisprotects against swelling and lysis

Despite the lack of a cell wall they Despite the lack of a cell wall they are able to survive in harsh are able to survive in harsh environments and elude the environments and elude the defenses of the human bodydefenses of the human body

L bacteria( discovered by Lister L bacteria( discovered by Lister Institute)Institute)

Some bacteria spontaneously lose Some bacteria spontaneously lose their ability to form the cell walltheir ability to form the cell wall

These are wall deficient strains – that These are wall deficient strains – that may lose their cell wall – sometimes may lose their cell wall – sometimes due to the treatment with antibioticsdue to the treatment with antibiotics

Mycobacterium paratuberculosis –is a Mycobacterium paratuberculosis –is a bacterium associated with chronic bacterium associated with chronic and debilitating Crohn’s diseaseand debilitating Crohn’s disease

Archaeal Cell wallsArchaeal Cell walls

Lack peptidoglycanLack peptidoglycanCan be composed of proteins, Can be composed of proteins,

glycoproteins, or polysaccharidesglycoproteins, or polysaccharidesHyperthermophiles – these are Hyperthermophiles – these are

extremophiles that can withstand extremophiles that can withstand temperatures above boiling despite temperatures above boiling despite the lack of a Peptidoglycan cell wallthe lack of a Peptidoglycan cell wall

S layersS layers

S-layersS-layers Regularly structured layers of protein or Regularly structured layers of protein or

glycoproteinglycoprotein Common among Archaea, where they may be Common among Archaea, where they may be

the only structure outside the plasma the only structure outside the plasma membranemembrane

In some gram-positive bacteria, the S-layer is In some gram-positive bacteria, the S-layer is external to the murein wallexternal to the murein wall

In gram-negative bacteria, it is external to the In gram-negative bacteria, it is external to the outer membraneouter membrane

In both the S-layer is several molecules thickIn both the S-layer is several molecules thick

S-layersS-layers

Basically protein molecules with Basically protein molecules with carbohydrates attachedcarbohydrates attached

Resistant to proteolytic enzymes and Resistant to proteolytic enzymes and protein denaturing agentsprotein denaturing agents

In the intestinal parasite, In the intestinal parasite, Campylobacter jejuni Campylobacter jejuni protects protects against phagocytosisagainst phagocytosis

These S layers protect against These S layers protect against invasion from bacteriophagesinvasion from bacteriophages

S- layer of ArchaeanS- layer of Archaean

Functions of capsules, slime Functions of capsules, slime layers, and S layerslayers, and S layers

Protection from host defenses (e.g., phagocytosis)

Protection from harsh environmental conditions (e.g., desiccation)

Attachment to surfaces Protection from viral infection or Protection from viral infection or

predation by bacteriapredation by bacteria Protection from chemicals in Protection from chemicals in

environment (e.g., detergents)environment (e.g., detergents) Motility of gliding bacteriaMotility of gliding bacteria Protection against osmotic stressProtection against osmotic stress

Additional External Additional External Characteristics CharacteristicsCharacteristics Characteristics

Layers of material lying outside the Layers of material lying outside the cell wallcell wallCapsulesCapsules

usually composed of polysaccharidesusually composed of polysaccharideswell organized and not easily removed from well organized and not easily removed from

cellcellSlime layersSlime layers

similar to capsules except diffuse, similar to capsules except diffuse, unorganized and easily removedunorganized and easily removed

CapsulesCapsules

Capsules and slime layersCapsules and slime layers

Nutritional environment may Nutritional environment may influence the formation of the influence the formation of the capsule or slime layercapsule or slime layer

Haemophilus influenzaHaemophilus influenza and and Streptococcus pneumoniaeStreptococcus pneumoniae are are pathogenic with capsules due to their pathogenic with capsules due to their ability to avoid phagocytic cells of ability to avoid phagocytic cells of the immune systemthe immune system

Slime layersSlime layers

This outer covering is a major This outer covering is a major determinant in the colonization of a determinant in the colonization of a nicheniche

Such is the case with the bacterium, Such is the case with the bacterium, Streptococcus mutans, Streptococcus mutans, this allows it this allows it to colonize the nooks and crannies of to colonize the nooks and crannies of your teeth to cause dental caries and your teeth to cause dental caries and participate in a biofilm on the surface participate in a biofilm on the surface of teethof teeth

GlycocalyxGlycocalyx

GlycocalyxGlycocalyxNetwork of polysaccharides extending Network of polysaccharides extending

from the surface of the cellfrom the surface of the cellA capsule or slime layer composed of A capsule or slime layer composed of

polysaccharides can also be referred to polysaccharides can also be referred to as a glycocalyxas a glycocalyx

The Nature of MembranesThe Nature of Membranes

Membranes are an absolute Membranes are an absolute requirement for all living organismsrequirement for all living organisms

Plasma membrane Plasma membrane encompasses the encompasses the cytoplasmcytoplasm

Some procaryotes also have internal Some procaryotes also have internal membrane systemsmembrane systems

Functions of Cell Functions of Cell MembranesMembranes

Separation of cell from its environmentSeparation of cell from its environment Selectively permeable barrierSelectively permeable barrier

some molecules are allowed to pass into or out some molecules are allowed to pass into or out of the cellof the cell

transport systems aid in movement of transport systems aid in movement of moleculesmolecules

Location of crucial metabolic processesLocation of crucial metabolic processes Detection of and response to chemicals in Detection of and response to chemicals in

surroundings with the aid of special surroundings with the aid of special receptor molecules in the membranereceptor molecules in the membrane

Lipid BilayerLipid Bilayer

Polar endsPolar ends interact with interact with

waterwaterhydrophilichydrophilic

Nonpolar endsNonpolar ends insoluble in waterinsoluble in waterhydrophobichydrophobic

Lipids and ProteinsLipids and Proteins Contains Phospholipids and Contains Phospholipids and

proteinsproteins lipids usually form a lipids usually form a

bilayerbilayer proteins are embedded in proteins are embedded in

or associated with lipidsor associated with lipids Highly organized, asymmetric, Highly organized, asymmetric,

flexible, and dynamicflexible, and dynamic Bacterial cell membranes are Bacterial cell membranes are

more similar to eukaryotes more similar to eukaryotes than Archaeathan Archaea

They have ester linkages like They have ester linkages like eukaryotes in their eukaryotes in their phopholipidsphopholipids

Cell Membrane ResearchCell Membrane Research http://www.rxpgnews.com/article_4916.shtmlhttp://www.rxpgnews.com/article_4916.shtml ““The discovery also demonstrated that current textbooks The discovery also demonstrated that current textbooks

use the wrong type of bacterium as a model to explain a use the wrong type of bacterium as a model to explain a critical biochemical step that most disease-causing bacteria critical biochemical step that most disease-causing bacteria use to make their membranes, according to Charles Rock, use to make their membranes, according to Charles Rock, Ph.D., a member of the St. Jude Department of Infectious Ph.D., a member of the St. Jude Department of Infectious Diseases and senior author of the paper. As bacteria grow Diseases and senior author of the paper. As bacteria grow in size or divide, they must make additional membrane in size or divide, they must make additional membrane using a series of biochemical reactions. The first step in this using a series of biochemical reactions. The first step in this process is the transfer of a fatty acid to a molecule called process is the transfer of a fatty acid to a molecule called G3P. Bacteria then convert this molecule into a variety of G3P. Bacteria then convert this molecule into a variety of other molecules called phospholipids, which are the other molecules called phospholipids, which are the building blocks of membranes.”building blocks of membranes.”

Archaeal Cell MembranesArchaeal Cell Membranes

Contain unique lipids call isoprenoids. Contain unique lipids call isoprenoids. These are arranged in bilayersThese are arranged in bilayers

These are also linked to glycerol by an These are also linked to glycerol by an ether linkage instead of an ether linkageether linkage instead of an ether linkage

Some membranes are single layers – The Some membranes are single layers – The molecules are longer than phospholipids molecules are longer than phospholipids and have glycerol molecules at both endsand have glycerol molecules at both ends

http://www.sciencemag.org/cgi/content/abhttp://www.sciencemag.org/cgi/content/abstract/293/5527/92stract/293/5527/92

Cytoplasmic MatrixCytoplasmic Matrix

Substance between Substance between membrane and membrane and nucleoidnucleoid

Packed with Packed with ribosomes and ribosomes and inclusion bodiesinclusion bodies

Highly organized Highly organized with respect to with respect to protein locationprotein location

Specialized Internal Specialized Internal MembranesMembranes

Complex in-foldings of the plasma Complex in-foldings of the plasma membranemembraneobserved in many photosynthetic observed in many photosynthetic

bacteria and in procaryotes with high bacteria and in procaryotes with high respiratory activityrespiratory activity

may be aggregates of spherical vesicles, may be aggregates of spherical vesicles, flattened vesicles, or tubular flattened vesicles, or tubular membranesmembranes

Internal MembranesInternal Membranes

MesosomesMesosomesMay be invaginations of the plasma May be invaginations of the plasma

membranemembranepossible rolespossible roles

cell wall formation during cell divisioncell wall formation during cell division chromosome replication and distributionchromosome replication and distribution secretory processessecretory processes

May be artifacts of chemical fixation May be artifacts of chemical fixation processprocess

The Nucleoid RegionThe Nucleoid Region Irregularly shaped Irregularly shaped

regionregion Location of chromosomeLocation of chromosome

usually 1/cellusually 1/cell Not membrane-boundNot membrane-bound The nucleoid region has

been isolated and analyzed

• 60% DNA, 30% RNA, and 10% protein. It has been stained with Feulgen that demonstrates the presence of DNA

Nucleoid characteristicsNucleoid characteristics If stretched out the DNA of E. coli would If stretched out the DNA of E. coli would

be 1000x times the length of the cellbe 1000x times the length of the cell The folding of the DNA – its packaging The folding of the DNA – its packaging

forms the nucleoid( the result of proteins )forms the nucleoid( the result of proteins ) When bacterial cells undergo lysis – and When bacterial cells undergo lysis – and

the interior contents of the cell are the interior contents of the cell are released, the viscosity or thickness is due released, the viscosity or thickness is due to the nucleoidto the nucleoid

Due to the density of the nucleoid, the Due to the density of the nucleoid, the transcription of DNA takes place at the transcription of DNA takes place at the nucleoid and cytoplasmic interfacenucleoid and cytoplasmic interface

Bacterial chromosomesBacterial chromosomes

The most common form of a bacterial The most common form of a bacterial chromosome is a ds circular chromosome is a ds circular chromosomechromosome

ExceptionsExceptionsSome procaryotes have Some procaryotes have > 1 > 1

chromosomechromosomeSome procaryotes have chromosomes Some procaryotes have chromosomes

composed of linear double-stranded composed of linear double-stranded DNADNA

A few genera have membrane-A few genera have membrane-delimited nucleoidsdelimited nucleoids

Bacterial chromosomesBacterial chromosomes

Circular chromosomesCircular chromosomes

The circular chromosomes have ends The circular chromosomes have ends that are protected due to the that are protected due to the structurestructure

In the linear chromosomes of In the linear chromosomes of prokaryotes the ends are protected prokaryotes the ends are protected by hairpins or by binding proteinsby hairpins or by binding proteins

Eyeing Bacterial GenomesEyeing Bacterial Genomes

Bacterial chromosomes can range Bacterial chromosomes can range from 580,000 base pairs to 10 million from 580,000 base pairs to 10 million base pairsbase pairs

The cholera bacterium has two The cholera bacterium has two dissimilar chromosomes while dissimilar chromosomes while nitrogen fixing bacterium have three( nitrogen fixing bacterium have three( it is somewhat of a mystery as to the it is somewhat of a mystery as to the apportioning of these chromosomes apportioning of these chromosomes during cell division)during cell division)

Eyeing Bacterial GenomesEyeing Bacterial Genomes All species of All species of BorreliaBorrelia have have

linear chromosomes linear chromosomes ranging in size from ranging in size from 900,000 to 920,000 base 900,000 to 920,000 base pairs, with an pairs, with an accompaniment of circular accompaniment of circular and linear plasmids (some and linear plasmids (some species contain up to 20 species contain up to 20 different plasmids). different plasmids).

Between the linear Between the linear chromosome and array of chromosome and array of plasmids there is a high plasmids there is a high degree of redundancy in degree of redundancy in the genetic sequence. the genetic sequence.

Borrelia burgdorferi

PlasmidsPlasmids Antibiotic-resistance genes Antibiotics production

genes Heavy Metal resistance

genes Virulence genes Tumorigenicity (in plants) Fertility (transfer) genes Toxin production Restriction / Modification Metabolism of

hydrocarbons

RibosomesRibosomes Complex structures Complex structures

consisting of protein consisting of protein and RNAand RNA

Sites of protein Sites of protein synthesissynthesis

Smaller than Smaller than eucaryotic ribosomeseucaryotic ribosomes procaryotic ribosomes procaryotic ribosomes

70S 70S eucaryotic ribosomes eucaryotic ribosomes

80S80S S = Svedburg unitS = Svedburg unit

Bacterial RibosomeBacterial Ribosome

Small Sub UnitSmall Sub Unit 30S30S 16S RNA16S RNA 21 proteins21 proteins

Large SubunitLarge Subunit 50S50S 23S & 5S RNAs23S & 5S RNAs 31 proteins31 proteins

InclusionsInclusions

Granules of organic or inorganic material. Used for storage of a variety of substances

like phosphates and glycogen. Most of these inclusion bodies are free in the cytoplasm.

Some inclusion bodies are enclosed by a thin membrane. Examples of these include carboxysomes and gas vacuoles.

The number of inclusion bodies varies with the nutritional status of the cells

PHBPHB

Poly- hydroxybutyrate ( PHB) contains hydroxybutyrate molecules joined by ester bonds between the carboxyl and hydroxyl of adjacent molecules. These are common in purple sulfur bacteria and stain with Sudan black for light microscopy. These granules serve as storage reservoirs for glycogen and sugars necessary for energy and biosynthesis.

Inclusions in CyanobacteriaInclusions in Cyanobacteria Cyanophycin granules are found in

Cyanobacteria. They are large inclusion bodies composed of polypeptides comprised of arginine and aspartic acid. These store additional nitrogen for the bacteria.

Cyanobacteria, thiobacilli, and nitrifying bacteria, organisms that reduce CO2 in order to produce carbohydrates, possess carboxysomes containing an enzyme used for CO2 fixation.

EnterosomesEnterosomes In In Salmonella and E. coli Salmonella and E. coli have internal have internal

structures similar to carboxysomesstructures similar to carboxysomes Enterosomes contain enzymes required Enterosomes contain enzymes required

for the metabolism of certain moleculesfor the metabolism of certain molecules The existence of these molecules may be The existence of these molecules may be

due to the necessity of dealing with toxic due to the necessity of dealing with toxic moleculesmolecules

Propanediol is a metabolite of fucose which Propanediol is a metabolite of fucose which is a sugar found on the intestinal wall of is a sugar found on the intestinal wall of mammals that that can be degraded by mammals that that can be degraded by intestinal bacteria – This is one of the intestinal bacteria – This is one of the molecules metabolized in enterosomesmolecules metabolized in enterosomes

Gas VacuolesGas Vacuoles• Purple and green

photosynthetic bacteria as well as some other aquatic bacteria contain gas vacuoles. These are aggregates of hollow protein cylinders called gas vesicles that are permeable to atmospheric gas, enabling the organism to regulate buoyancy. Bacteria are able to regulate the depth at which they float to regulate photosynthetic activity

VolutinVolutin Some bacteria produce

inorganic inclusion bodies in their cytoplasm, including volutin granules that store phosphate and sulfur granules that store sulfur. Volutin is a source of phosphate for DNA. Sulfur is used by purple photosynthetic bacteria that use hydrogen sulfide as a photosynthetic electron donor.

MagnetosomesMagnetosomes• Some motile aquatic

bacteria are able to orient themselves by responding to the magnetic fields of the earth because they possess magnetosomes, membrane-bound crystals of magnetite or other iron-containing substances that function as tiny magnets.

MagnetosomesMagnetosomes

Movement of bacteria in a Movement of bacteria in a magnetic fieldmagnetic field

External StructuresExternal Structures

FimbriaeFimbriaePiliPiliFlagellaFlagella

PiliPili

• Pili are appendages that are larger than fimbriae. Their presence is determined by genes on plasmids called sex factors. These structures function in conjugation which is a genetic exchange occurring in bacteria with these appendages

FimbriaeFimbriae• Fimbriae are thin, hair-

lie projections extending from the cell wall in Gram – bacteria. They are composed of helical protein units and designed for attachment to the host cell membranes( mucous). They also may contribute to types of movement in some bacteria.

Neisseria gonorrhea

Adhesion and colonizationAdhesion and colonization

An essential step in the successful An essential step in the successful colonization and production of colonization and production of disease is their ability to adhere.disease is their ability to adhere.

Bacterial molecules utilized for Bacterial molecules utilized for adhesion belong to a class called adhesion belong to a class called adhesinsadhesins

AdhesinsAdhesins are proteins that are found are proteins that are found in folds on the bacterial surface or on in folds on the bacterial surface or on a pilus or a fimbriaea pilus or a fimbriae

Example of an Adhesin in a Example of an Adhesin in a PathogenPathogen

E. coliE. coli uses an adhesion on pili to uses an adhesion on pili to bind to the lining of the urinary tract bind to the lining of the urinary tract to cause infection of the kidneyto cause infection of the kidney

The adhesin is associated with a P The adhesin is associated with a P pilus regarded as vital to the pilus regarded as vital to the adhesion processadhesion process

Receptors on the host lining of the Receptors on the host lining of the urinary tract are used for this urinary tract are used for this adhesive phenomenonadhesive phenomenon

Flagella MotilityFlagella Motility

http://www-micro.msb.le.ac.uk/video/motility.html

Arrangement of flagellaArrangement of flagella

monotrichousmonotrichous – one flagellum – one flagellumpolar flagellumpolar flagellum – flagellum at end of – flagellum at end of

cellcellamphitrichousamphitrichous – one flagellum at each – one flagellum at each

end of cellend of cell lophotrichouslophotrichous – cluster of flagella at – cluster of flagella at

one or both endsone or both endsperitrichousperitrichous – spread over entire – spread over entire

surface of cellsurface of cell

Arrangement of FlagellaArrangement of Flagella

The three parts of the The three parts of the flagellumflagellum

3 parts3 partsfilamentfilamentbasal bodybasal bodyhookhook

Structure of Bacterial Structure of Bacterial FlagellaFlagella

The filamentThe filament

Hollow, rigid cylinderHollow, rigid cylinderComposed of the protein Composed of the protein flagellinflagellinSome prokaryotes have a sheath Some prokaryotes have a sheath

around filamentaround filamentFlagellins are highly antigenic. They Flagellins are highly antigenic. They

are extremely rigid in natureare extremely rigid in nature

The hook and basal bodyThe hook and basal body

HookHooklinks filament to basal bodylinks filament to basal body

The hook is a short-curved structure slightly The hook is a short-curved structure slightly larger than in diameter than the filamentlarger than in diameter than the filament

The hook is curved The hook is curved

Basal bodyBasal body Series of rings that drive flagellar motorSeries of rings that drive flagellar motor It is composed of 15 proteins thatIt is composed of 15 proteins that

aggregate to form a rod to which four rings are aggregate to form a rod to which four rings are attachedattached

Gram positive and gram negative bacteria have Gram positive and gram negative bacteria have different attachmentsdifferent attachments

Flagellar complexityFlagellar complexity

Gram Positive and Gram Gram Positive and Gram NegativeNegative

Differ in the construction of their rings or Differ in the construction of their rings or basal bodybasal body

Gram positive have an S and M ring- an Gram positive have an S and M ring- an inner ring connected to the plasma inner ring connected to the plasma membrane and an outer ring connected to membrane and an outer ring connected to the peptidoglycan cell wallthe peptidoglycan cell wall

Gram negative have an S and M and an L Gram negative have an S and M and an L and P, The L associates with the LPS anda and P, The L associates with the LPS anda the P associates with the peptidoglycanthe P associates with the peptidoglycan

Flagellar SynthesisFlagellar Synthesis

An example of An example of self-assemblyself-assemblyComplex process involving many Complex process involving many

genes and gene productsgenes and gene productsNew molecules of flagellin are New molecules of flagellin are

transported through the hollow transported through the hollow filamentfilament

Growth is from tip, not baseGrowth is from tip, not base

Flagellar SynthesisFlagellar Synthesis

Flagellar MotionFlagellar Motion

flagellum rotates like a propellerflagellum rotates like a propeller in general, counterclockwise rotation in general, counterclockwise rotation

causes forward motion (causes forward motion (runrun)) in general, clockwise rotation disrupts in general, clockwise rotation disrupts

run causing a run causing a tumble tumble ((twiddletwiddle))

Tumble and RunTumble and Run

Other Types of MotilityOther Types of Motility

SpirochetesSpirochetesaxial filamentsaxial filaments cause flexing and cause flexing and

spinning movementspinning movementGliding motilityGliding motility

cells coast along solid surfacescells coast along solid surfacesno visible motility structure has been no visible motility structure has been

identifiedidentified

ChemotaxisChemotaxis

Movement towards a chemical Movement towards a chemical attractant or away from a chemical attractant or away from a chemical repellantrepellant

Concentrations of chemoattractants Concentrations of chemoattractants and chemorepellants detected by and chemorepellants detected by chemoreceptorschemoreceptors on surfaces of cells on surfaces of cells

ChemotaxisChemotaxis

Negative chemotaxis – Increasing concentrations of acetate are applied to disk – see the increasing clear zone from right to left – suggesting movement away

Positive chemotaxis – Left ring is caused by bacteria consuming the amino acid serine. The right ring a less attractive aspartate attracts fewer bacteria

Traveling toward and Traveling toward and AttractantAttractant

Caused by lowering Caused by lowering the frequency of the frequency of tumblestumbles

Traveling away Traveling away involves similar but involves similar but opposite responsesopposite responses

ChemoreceptorsChemoreceptors

Bacteria detect attractants and Bacteria detect attractants and repellants at the molecular levelrepellants at the molecular level

The chemosensing system consists The chemosensing system consists of proteins that may collect in the of proteins that may collect in the periplasmic space or the plasma periplasmic space or the plasma membranemembrane

The receptors may be organized in The receptors may be organized in patches on the membranepatches on the membrane

E. coliE. coli

Has four rceptors each of which Has four rceptors each of which recognize serine, aspartate, maltose, recognize serine, aspartate, maltose, ribose, galactose , and dipeptides.ribose, galactose , and dipeptides.

These chemoreceptors are calledThese chemoreceptors are called

MCP’s methyl accepting proteinsMCP’s methyl accepting proteinsThese are found on the ends of the These are found on the ends of the

rod shaped bacillusrod shaped bacillus

Complexity of reaction to Complexity of reaction to stimulistimuli

Receptor and molecule bind causing Receptor and molecule bind causing conformational changes in the receptor conformational changes in the receptor that are transmitted through the that are transmitted through the membranemembrane

The CheA protein is phosphorylated using The CheA protein is phosphorylated using ATP ATP

This provides a phosphate for the Che YThis provides a phosphate for the Che Y The Che Y then interacts with FliM that is The Che Y then interacts with FliM that is

at the base of the flagella and regulates at the base of the flagella and regulates flagellar motionflagellar motion

Bacterial EndosporesBacterial Endospores

formed by some bacteriaformed by some bacteriadormantdormant resistant to numerous environmental resistant to numerous environmental

conditionsconditionsheatheatradiationradiationchemicalschemicalsdesiccationdesiccation

Position of endosporePosition of endospore

Resistance of endosporeResistance of endospore

Calcium (complexed with Calcium (complexed with dipicolinic dipicolinic acidacid))

Acid-soluble, DNA-binding proteinsAcid-soluble, DNA-binding proteinsDehydrated coreDehydrated coreSpore coatSpore coatDNA repair enzymesDNA repair enzymes

Electron Micrograph of Electron Micrograph of endosporeendospore

CW = Vegetative CW = Vegetative cell wallcell wall

CP= Spore CoatCP= Spore Coat SC= Spore CortexSC= Spore Cortex EX= ExosporiumEX= Exosporium

SporogenesisSporogenesis

Normally commences when growth Normally commences when growth ceases because of lack of nutrientsceases because of lack of nutrients

Complex multistage processComplex multistage process

Formation of the Vegetative Formation of the Vegetative Cell- Sporulation or Cell- Sporulation or

SporogenesisSporogenesis Complex, Complex,

multistage processmultistage process Commences in Commences in

response to response to environmental environmental conditions such as conditions such as a lack of nutrientsa lack of nutrients

StepsSteps

The nuclear material formsThe nuclear material forms Inward folding of the cell membrane to Inward folding of the cell membrane to

enclose part of the DNA and produce the enclose part of the DNA and produce the forespore septumforespore septum

The membrane continues to grow and The membrane continues to grow and engulfs the immature spore in a second engulfs the immature spore in a second membrane.membrane.

The cortex is then laid down in the space The cortex is then laid down in the space between the two membranes and dipocolinic between the two membranes and dipocolinic acid and Calcium ions are accumulatedacid and Calcium ions are accumulated

Sporulation continuedSporulation continued

Protein coats are then formed around Protein coats are then formed around the cortexthe cortex

Maturation of the spore occursMaturation of the spore occurs

Steps in ActivationSteps in Activation ActivationActivation

prepares spores for germinationprepares spores for germination often results from treatments like heatingoften results from treatments like heating

GerminationGermination spore swellingspore swelling rupture of absorption of spore coatrupture of absorption of spore coat loss of resistanceloss of resistance increased metabolic activityincreased metabolic activity

OutgrowthOutgrowth emergence of vegetative cellemergence of vegetative cell

Protein Secretion Systems in E. Protein Secretion Systems in E. colicoli

Protein Secretion in ProkaryotesProtein Secretion in Prokaryotesnumerous protein secretion numerous protein secretion

pathways have been identifiedpathways have been identified four major pathways are:four major pathways are:

Sec-dependent pathwaySec-dependent pathwaytype II pathwaytype II pathwaytype I (ABC) protein secretion pathwaytype I (ABC) protein secretion pathwaytype III protein secretion pathwaytype III protein secretion pathway

Protein Secretion – Sec Protein Secretion – Sec DependentDependent

Sec-Dependent PathwaySec-Dependent Pathway Also called Also called general secretion pathwaygeneral secretion pathway Translocates proteins from cytoplasm across or Translocates proteins from cytoplasm across or

into plasma membraneinto plasma membrane Secreted proteins synthesized as preproteins Secreted proteins synthesized as preproteins

having amino-terminal having amino-terminal signal peptidesignal peptide signal peptide delays protein foldingsignal peptide delays protein folding chaperone proteins keep preproteins unfoldedchaperone proteins keep preproteins unfolded

TransloconTranslocon transfers protein and removes signal transfers protein and removes signal peptidepeptide

E. Coli and Sec Dependent E. Coli and Sec Dependent PathwayPathway

In E. coli the chaperones uesed for In E. coli the chaperones uesed for transport are Sec B and the Signal transport are Sec B and the Signal Recognition particle SRP.Recognition particle SRP.

Sec B is found in Gram negative Sec B is found in Gram negative bacteria and SRP is found in all bacteria and SRP is found in all prokaryotesprokaryotes

Steps in protein secretionSteps in protein secretion

Sec B binds to Sec A portion of the Sec B binds to Sec A portion of the translocon, which is the transport translocon, which is the transport machinerymachinery

The preprotein is transferred to SecAThe preprotein is transferred to SecAThe protein can be released by The protein can be released by

hydrolysis of GTPhydrolysis of GTPAfter this has occurred the protein is After this has occurred the protein is

transferred through the membranetransferred through the membrane

TransloconTranslocon The bacterial trnaslocaon si composed of a The bacterial trnaslocaon si composed of a

membrane protein complex called SecYEG, membrane protein complex called SecYEG, SecA and other proteinsSecA and other proteins

It is believed that his complex forms a It is believed that his complex forms a channel in the membrane through which channel in the membrane through which the protein passesthe protein passes

Energy is required for this process in the Energy is required for this process in the form of ATP hydrolysis coupled with proton form of ATP hydrolysis coupled with proton motive force.( Archaea do not possess this motive force.( Archaea do not possess this mechanism)mechanism)

Structure of the Sec Structure of the Sec Dependent PathwayDependent Pathway

Sec Dependent Sec Dependent PathwayPathway

ABC TransportersABC Transporters

Also called Also called ABC protein secretion pathwayABC protein secretion pathway Transports proteins from cytoplasm across Transports proteins from cytoplasm across

both plasma membrane and outer both plasma membrane and outer membranemembrane

Secreted proteins have C-terminal Secreted proteins have C-terminal secretion signalssecretion signals

Proteins that comprise type I systems form Proteins that comprise type I systems form channels through membraneschannels through membranes

Translocation driven by both ATP Translocation driven by both ATP hydrolysis and proton motive forcehydrolysis and proton motive force

Type IIType II

Transports proteins from periplasmic across outer membrane

Present in Pseudomonas aeruginosa and Vibrio cholera

Observed in some gram-negative bacteria, including some pathogens

Complex systems consisting of up to 12-14 proteinsmost are integral membrane proteins

ABC TransportersABC Transporters

Type IType I

Type III and SecretionType III and Secretion

Secretes virulence factors of gram-Secretes virulence factors of gram-negative bacteria from cytoplasm, negative bacteria from cytoplasm, across both plasma membrane and across both plasma membrane and outer membrane, and into host cellouter membrane, and into host cell

Some type III secretion machinery is Some type III secretion machinery is needle-shapedneedle-shapedsecreted proteins thought to move secreted proteins thought to move

through a translocation channelthrough a translocation channel

OccurrenceOccurrence

Found in Found in Salmonella, Pseudomonas, Salmonella, Pseudomonas, Yersinia, ShigellaYersinia, Shigella, and , and E. coliE. coli

Contact between the bactgeria and Contact between the bactgeria and the host cells simtulates the processthe host cells simtulates the process

Low calcium levels may be required Low calcium levels may be required for secretionfor secretion

Type III and virulence factorsType III and virulence factors

Type III Secretion Type III Secretion PathwayPathway

Four different types of Four different types of proteinsproteins

The secretory portion, The secretory portion, the regulators, the the regulators, the proteins that aid in the proteins that aid in the insertion of secreted insertion of secreted proteins, and effectors proteins, and effectors that alter host that alter host functionfunction

Examples of Type IIIExamples of Type III

CytotoxinsCytotoxinsPhagocytosis inhibitorsPhagocytosis inhibitorsStimulators for reorganization of the Stimulators for reorganization of the

cytoskeletoncytoskeletonApoptosis promotersApoptosis promoters