Prokaryotic ProfilesThe Bacteria and Archaea

Chapter 3

Prokaryotic Form and Function

Prokaryotes can be distinguished from eukaryotes by:

the way their DNA is packaged (lack of nucleus and histones)

the makeup of their cell wall (peptidoglycan and other unique chemicals)

their internal structure (lack of membrane-bounded organelles)

The Structure of the Prokaryotic Cell

All bacterial cells possess:

- a cell membrane

- cytoplasm

- ribosomes

- a cytoskeleton

- one (or a few) chromosome(s)

Most bacterial cells possess:

- a cell wall

- a surface coating called a glycocalyx

The Structure of the Prokaryotic Cell

Some but not all bacterial cells possess:

- flagella, pili, and fimbriae

- an outer membrane

- plasmids

- inclusions

- Endospores (or simply spores)

- intracellular membranes

Prokaryotic Cell

Cell (cytoplasmic) membrane—A thin sheet of lipidand protein that surrounds the cytoplasm and controlsthe flow of materials into and out of the cell pool.

In All Bacteria

Cytoplasm—Water-basedsolution filling the entire cell.

Ribosomes—Tiny particlescomposed of protein and RNAthat are the sites of proteinsynthesis.

Actin cytoskeleton—Long fibersof proteins that encircle the celljust inside the cell membrane andcontribute to the shape of the cell.

Bacterial chromosome or nucleoid—Composed ofcondensed DNA molecules. DNA directs all geneticsand heredity of the cell and codes for all proteins. Fimbriae—Fine, hairlike

bristles extending from thecell surface that help inadhesion to othercells and surfaces.

Cell wall—A semirigid casingthat provides structural supportand shape for the cell.

Outer membrane—Extramembrane similar to cellmembrane but alsocontaining popolysaccharide. Controls flow of materials,and portions of it are toxicto mammals when released.

In Some Bacteria

Inclusion/Granule—Stored nutrients suchas fat, phosphate, or glycogen deposited indense crystals or particles that can betapped into when needed.

Flagellum—Specialized appendage attachedto the cell by a basal body that holds a long,rotating filament. The movement pushes thecell forward and provides motility.

Pilus—An elongated, hollowappendage used in the transferof DNA to other cells.

Plasmid—Double-stranded DNA circlecontaining extra genes.

Capsule (tan coating)—A coating or layer ofmolecules external to the cell wall. It servesprotective, adhesive, and receptor functions.It may fit tightly or be very loose and diffuse.Also called slime layer and glycocalyx.

Endospore (not shown)—Dormant body formed withinsome bacteria that allows for their survival in adverse conditions.

Intracellular membranes(not shown)

In Some Bacteria (not shown)

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Prokaryotic Shapes and Arrangements

Most prokaryotes exist as unicellular organism

sometimes they can act as a group

colonies or in biofilms

On average, prokaryotic cells are 1 μm (microns

can range from 0.05 -0.2 μm (“nanobes”) to 750 μm

Cells of one species may vary in shape and size

this is pleomorphism caused by variations in cell wall structure

Arrangement of Cocci Resulting from Different Planes of Cell Division

Division inone plane

Diplococcus(two cells)

Streptococcus(variable numberof coci in chains)

Division in twoperpendicular

planes

Tetrad (cocci inpackets off our)

Sarcina(packet of8–64 cells)

Division inseveral planes

Irregular clusters(number of cells varies)

Staphylococci andMicrococci`

(c)(b)(a)

External Structures

Filament

Outermembrane

Cellwall

Cellmembrane

(b)(a)

Basalbody Rod

Hook

Rings

Noreen R. Francis, Gina E. Sosinsky, Dennis Thomas and David J. DeRosier, “Isolation, Characterization and Structure of Bacterial Flagellar Motors Containing the Switch Complex,” Journal of Molecular Biology, Vol 235, Issue 4, 27 January 1994, Pages 1261–1270.

Flagella – The Prokaryotic Propellers

Bacterial locomotion

Three distinct parts

Comprised of many proteins

360o rotation

Flagellar Arrangements

Monotrichous: single flagellum

Lophotrichous: small bunches or tufts of flagella

Amphitrichous: flagella at both poles of the cell

Peritrichous: flagella dispersed randomly over the surface of the cell

Bacterial Movement

Bacteria move in response to chemical signals (chemotaxis)

Receptors bind extracellular molecules, which triggers flagellum to rotate

Runs: smooth linear movement toward a stimulus

Tumbles: flagellar rotation reverses, causing the cell to stop and change its course

Straight Tumble

(a) General motility of a singular flagellum

Straight

(b) Peritrichous motility

Tumble

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Prokaryotic Appendages

Fimbriae: used for attachment

Pili: used for attachment and genetic exchange during conjugation

Fimbriae

Pili

Glycocalyx

First colonistsOrganic surfacecoating

Surface

(a)

Glycocalyxslime

Cathetersurface

Cellcluster

(b)

Cells stick tocoating.

As cells divide, theyform a dense matbound together bysticky extracellulardeposits.

Additional microbesare attracted todeveloping film andcreate a maturecommunity withcomplex function.

Glycocalyx

b: © Science VU–Charles W. Stratton/Visuals Unlimited

Composed of polysaccharides, proteins or both

Varies in thickness / take on form of slime layer or capsule

Used to avoid phagocytosis and for adhesion (biofilms)

CapsuleBound more tightly to the cell, denser and thicker than a slime layer

Visible by negative staining

Produces a sticky (mucoid) character to colonies

Encapsulated bacterial cells generally have greater pathogenicity

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Capsule

Cellbody

(a) (b)(a): © John D. Cunningham/Visuals Unlimited; b: Fig 1 from Brett J. Pellock, Max Teplitski, Ryan P. Boinay, W. Dietz Bauer, and Graham C. Walker,

“A LuxR Homolog Controls Production of Symbiotically Active Extracellular Polysaccharide II by Sinorhizobium meliloti.” Journal of Bacteriology, September15, 2002, Vol. 184, No. 18, p. 5067–5076. Reproduced with permission from American Society for Microbiology.

Cell Envelope

Lies outside of the cytoplasm

Composed of two or three basic layers:

cell membrane

cell wall

outer membrane in some bacteria

Peptidoglycan Cell Wall

Repeating framework of long glycan (sugar) chains cross-linked by short peptide(protein) fragments

Present in most bacteria

Provides strength to resist rupturing due to osmotic pressure

M

G

M

G

M

G

M

G

M

M

G

M

G

M

M

G

M

G

M

M

G

M

G

M

(a) The peptidoglycan can be seen as a crisscrossnetwork pattern similar to a chainlink fence.

(b) It contains alternating glycans (G and M) bound together inlong strands. The G stands for N-acetyl glucosamine, andthe M stands for N-acetyl muramic acid.

O

O

O

O

O

O

O

O

O

O

O

O

O

O

O

O

O

O

O

O

CC

HH3C H3C

M

O

OM

O

O

CC

H

L –

D –

L –

D –

–– glycine

– glycine ––

L –

D–

L –

D–

(c) A detailed view of the links between the muramic acids.Tetrapeptide chains branching off the muramic acids connectby interbridges also composed of amino acids. It is this linkagethat provides rigid yet flexible support to the cell and that maybe targeted by drugs like penicillin.

Tetr

apep

tide

OO

G

G

OO

G

GGlycan chains

Peptide cross-links

CH2OH

Interbridge

alanine

glutamate

lysine

alanine

alanine

glutamate

lysine

alanine

glycineglycine

glycine

CH2OH

Gram Positive Cell

Thick peptidoglycan

Teichoic acid and lipoteichoic acid

One membraneGram-Positive

Membrane proteins

Peptidoglycan

Cell membrane

Lipoteichoic acidWall teichoic acid

Envelope

Gram-Negative

Lipoproteins Porin proteins

Lipopolysaccharides

Phospholipids

Membraneprotein

Periplasmicspace

Membrane proteins

Outer membrane layer

Peptidoglycan

Cell membrane

Gram Negative Cell

Thin peptidoglycan

Lipopolysaccharide (endotoxin)

Two membranes / outer and inner

Porins

The Gram Stain: Developed by Hans Christian Gram (1884)

1. Crystal violetFirst, crystal violet is added to the cells in a smear. It stains themall the same purple color.

2. Gram’s iodineThen, the mordant, Gram’s iodine, is added. This is a stabilizer thatcauses the dye to form large complexes in the peptidoglycan meshworkof the cell wall. The thicker gram-positive cell walls are able to morefirmly trap the large complexes than those of the gram-negative cells.

3. AlcoholApplication of alcohol dissolves lipids in the outer membrane andremoves the dye from the peptidoglycan layer—only in thegram-negative cells.

4. Safranin (red dye)Because gram-negative bacteria are colorless after decolorization, theirpresence is demonstrated by applying the counterstain safranin in thefinal step.

Step Microscopic Appearance of Cell

Gram (+) Gram (–)

Chemical Reaction in Cell Wall(very magnified view)

Gram (+) Gram (–)

Both cell walls affix the dye

Dye complextrapped in wall

No effectof iodine

Crystals remainin cell wall

Outer membraneweakened; wall

loses dye

Red dye maskedby violet

Red dye stainsthe colorless cell

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© McGraw-Hill Companies, Inc.

Acid-Fast Bacteria

Mycobacterium sp. and Nocardia sp.

Cell wall contain a form of peptidoglycan but also Contain mycolic acid (a wax)

Gram-positive staining however

Use an acid-fast stain to stain these type of bacteria

Key to ID important pathogens causing:

tuberculosisleprosy

Cell-Wall Deficient Bacteria

Cell membrane stabilized by sterols, makes them resistant to lysisi.e. Mycoplasma pneumoniae

Other cell-wall deficient types called L forms are linked to infections

Cell wall(peptidoglycan)Cell membrane Peptidoglycan

lostProtoplast

Cell membrane

Outer membrane

Cell membrane

Spheroplast

Peptidoglycanlost

Outer membranePeptidoglycanCell membrane

Mutation or chemical treatment

Mutation or chemical treatment

Gram-Positive

Gram-Negative

(a)

(b)

Case FileArtwork

Lipopolysaccharide

Located in the outermost layer of the outer membrane (OM) in gram-negative bacteria

Lipid A is the factor which = endotoxin

Endotoxins cause fever and systemic shock

Case FileArtwork

Cytoplasmic Membrane

A lipid bilayer with proteins embedde

bacterial cell membranes contain primarily phospholipids (30%–40% of the membrane mass) and proteins (60%–70% of the membrane mass)

Provides a site for metabolic reaction

contains enzymes of respiration and ATP synthesis since prokaryotes lack mitochondria

Another major action of the cell membrane is to regulate the passage of nutrients into and out of the cell

Case FileArtwork

The Cytoplasm

70-80% water

Soluble proteins, salts, carbohydrates

Site of nearly all chemical reactions

Contains the DNA in the nucleoid

Case FileArtwork

Bacterial DNA

DNA of most bacteria exists in the form of a single circular bacterial chromosome

DNA is aggregated in a dense area of the cell called the nucleoid

Many bacteria contain other, nonessential pieces of DNA called plasmids

Largesubunit

(50S)

Smallsubunit

(30S)

Ribosome (70S)

Prokaryotic Ribosome

Two subunits(30S and 50S)

Total size is 70S (versus 80S in eukaryotes)

60% rRNA and 40% protein

Translates mRNA into proteins

Case FileArtwork

Inclusions or Granules:Storage Bodies

Non-membrane bound granules

Usually for storage of nutrients

The Cytoskeleton

Peptidoglycan layer determines shape of many bacteria

Others use protein fibers composed of actin and tubulin to alter cell shape

Case FileArtwork

Bacterial Endospores (or simply spore)

Dormant bodies

Heat resistance due to calcium and dipicolinicacid content

Cortex, spore coats protect against radiation and chemicals

Metabolically active vegetative cells can undergo sporulation

Sporulation is not a reproductive function for most bacteria

When spores of Clostridium sp. are embedded in a wound with dead tissue, they can germinate, grow, and release toxins

A Typical Sporulation Cycle in Bacillus Species

1

2

3

4

56

7

8

9

Spore coats

Chromosome

Cortex

Core of spore

Germination:spore swellsand releasesvegetative cell.

Frees pore isreleased with theloss of thesporangium.

ExosporiumSpore coatCortexCore

Matureendospore

Cortex andouter coat layersare deposited.

Cortex Early spore

ForesporeSporangium

Cell wall Cell membrane

Chromosome

Vegetative cell begins tobe depleted of nutrients.

Chromosome isduplicated andseparated.

Cell is septatedinto a sporangiumand forespore.

Sporangium engulfsforespore for furtherdevelopment.

Sporangium begins toactively synthesizespore layers aroundforespore.

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ArchaeaProkaryotic microorganisms

Many are found in extreme environments (i.e. psychrophiles)

Different from members of the domains Bacteria and Eukarya in terms of: cell structure, metabolism, genetics

Able to survive in extreme environments / metabolize ammonia, methane, hydrogen!

Table 3.2 Comparison of Three Cellular Domains

70S but structure is similarto 80S

Contains unique ribosomal RNAsignature sequences

+

Number of sequences sharedwith Eukarya

1

Protein synthesis similar to Eukarya

Presence of peptidoglycan in cell wall +

Fatty acids with ester linkages Long-chain, branchedhydrocarbons with ether linkages

Fatty acids with esterlinkages

+

Characteristic

Cell type

ChromosomesTypes of ribosomes

Cell membrane lipids

Sterols in membrane

Archaea

Prokaryotic

Single, circular

+

3

+

Bacteria

Prokaryotic

Single, or few circular70S

(some exceptions)

(all)

+

+

80SSeveral, linear

Eukaryotic

Eukarya

Classification Systems

For differentiating and identifying unknown microbial species i.e. Bergey’s Manual of Determinative Bacteriology(phenotypic data)

For studying prokaryotic relationships and originsi.e. Bergey’s Manual of Systematic Bacteriology (rRNAsequencing data)

Streptococcus pyogenesRosenbach 1884

Binomial name

S. pyogenesSpecies:

StreptococcusGenus:

StreptococcaceaeFamily:

LactobacillalesOrder:

CocciClass:

FirmicutesPhylum:

EubacteriaKingdom:

Scientific classification

Streptococcus pyogenesS. pyogenes bacteria at 900x magnification.

Case FileArtwork

Prokaryotic Classification Systems

Bergey’s Manual of Determinative Bacteriology organizes the prokaryotes into four major divisions based on cell wall structure:

Gracilicutes: gram-negative cell walls and thus are thin-skinned

Firmicutes: gram-positive cell walls that are thick and strong

Tenericutes: lack a cell wall and thus are soft

Mendosicutes: archaea with unusual cell walls

Species and Subspecies in Prokaryotes

Theoretically, a collection of bacterial cells, all of which share an overall similar pattern of traits and 70%–80% of their genes

Members of given species can show variations

subspecies, strain, or type are terms used to designate bacteria of the same species that have differing characteristics

serotype refers to representatives of a species that stimulate a distinct pattern of antibody (serum) responses in their hosts

Case FileArtwork

Concept Check

Which of the following structures are possessed by

some but not all prokaryotes?

A. Ribosomes

B. A cell membrane

C. One or more chromosomes

D. Flagella

Concept Check

Which of the following bacterial appendages is not used for attachment?

A. Slime layer

B. Flagellum

C. Pilus

D. Fimbriae

Concept CheckThe outer membrane contributes an extra barrier in gram-positive bacteria that makes them impervious to some antimicrobial chemicals, so they are generally more difficult to inhibit or kill than are gram-negative bacteria.

A. True

B. False

Concept Check

Where in a prokaryotic cell would you find the genetic material?

A. Nucleus

B. Nucleolus

C. Nucleocapsid

D. Nucleoid

Concept Check

Which of the follow do members of the domains Bacteria and Archaea both possess?

A. Linear DNA

B. A nucleus

C. 70S ribosomes

D. Fatty acids with ether linkages

Concept CheckYou identify a bacterium as gram-negative using the Gram stain method. In Bergey’s Manual of Determinative Bacteriology, this bacterium belongs to the:

A. Gracilicutes

B. Firmicutes

C. Tenericutes

D. Mendosicutes