Download Lecture 2 - Cell assembly

Chemicals needed for life
• Besides chemicals for metabolic energy,
microbes need other things for growth.
–
–
–
–
–
–
–
Carbon
Oxygen
Sulfur
Phosphorus – Arsenic can substitute (??)
Nitrogen
Iron
Trace metals (including Mo, Cu, Ni, Cd, etc.)
• LET’S PUT THIS TOGETHER INTO A
MICROBE….
Cell Composition
• 70-90% water
• Organic chemistry key to the construction of
cells is inherently linked to the properties of
water vs. organic compounds
• Consider 4 groups of monomers (a single,
repeated ‘building block’):
Polysaccharides
Lipids
Nucleic Acids
Proteins
Macromolecules
– Sugars
– Fatty Acids
– Nucleotides
– Amino Acids
Macromolecules
• Informational macromolecules: They carry
information because the sequence of
monomer building blocks is specific and
carries information = Nucleic Acids and
Proteins
• Non-informational macromolecules: The
sequence is highly repetitive and the
sequence has no function to carry
information
• composition and how exactly the sequences
are structures delineate different functionality
Small molecules present in a growing bacterial cell.
Monomers
Approximate ##
of kinds
Amino acids, their precursors and derivatives
120
Nucleotides, their precursors and derivatives
100
Fatty acids and their precursors
50
Sugars, carbohydrates and their precursors or derivatives
250
quinones, porphyrins, vitamins, coenzymes and prosthetic
300
groups and their precursors
Molecular composition of E. coli under conditions of balanced growth.
Molecule
Protein
Total RNA
DNA
Phospholipid
Lipopolysaccharide
Murein
Glycogen
Small molecules: precursors,
metabolites, vitamins, etc.
Inorganic ions
Total dry weight
Percentage
of dry
weight
55
20.5
3.1
9.1
3.4
2.5
2.5
2.9
1.0
100.0
Inorganic ions present in a growing bacterial cell.
Ion
Function
K+
Maintenance of ionic strength; cofactor for certain enzymes
NH4+
Principal form of inorganic N for assimilation
Ca++
Cofactor for certain enzymes
Fe++
Present in cytochromes and other metalloenzymes
Mg++
Cofactor for many enzymes; stabilization of outer membrane of Gramnegative bacteria
Mn++
Present in certain metalloenzymes
Co++
Trace element constituent of vitamin B12 and its coenzyme derivatives and
found in certain metalloenzymes
Cu++
Trace element present in certain metalloenzymes
Mo++
Trace element present in certain metalloenzymes
Ni++
Trace element present in certain metalloenzymes
Zn++
Trace element present in certain metalloenzymes
SO4--
Principal form of inorganic S for assimilation
PO4---
Principal form of P for assimilation and a participant in many metabolic
reactions
Cell Composition
• 70-90% water
• Organic chemistry key to the construction of
cells is inherently linked to the properties of
water vs. organic compounds
• Consider 4 groups of monomers (a single,
repeated ‘building block’):
Polysaccharides
Lipids
Nucleic Acids
Proteins
Macromolecules
– Sugars
– Fatty Acids
– Nucleotides
– Amino Acids
Construction, Part 1…
• Sugars (aka carbohydrates) can be linear or cyclic
(if >5 C)
• Sugars start out with 4,5,6, or 7 carbons:
• Pentoses (C5) are critical to DNA, RNA (form the
‘backbone’)
– Hexoses (C6) are crucial to cell walls
• Polysaccharides contain hundreds of sugars or
more held together with glycosidic bonds with
either a or b orientations
• Cn(H2O)n-1 where n is typically 200-2500
a
b
Polysaccharides:
•
•
•
•
Glycogen – C and energy storage
Starches – C and energy storage (a poly)
Cellulose – cellular wall material (b poly)
Extracellular polysaccharides (aka
glycoproteins or glycolipids) - pathogenic
component of some cells, also useful for
attachment and solubilization
Construction, Part 2
• Fatty Acids – long chains of C (aliphatic)
• Lipids are made of fatty acids put together
to form hydrophobic and hydrophilic end
The chemical characteristics
of the fatty acids and
subsequently the lipids make
them ideal for membranes
Cell Composition
• 70-90% water
• Organic chemistry key to the construction of
cells is inherently linked to the properties of
water vs. organic compounds
• Consider 4 groups of monomers (a single,
repeated ‘building block’):
Polysaccharides
Lipids
Nucleic Acids
Proteins
Macromolecules
– Sugars
– Fatty Acids
– Nucleotides
– Amino Acids
Construction, Part 3
• Bases – Two types:
Pyrimidine
Purine
• Derivatives
Cytosine, C
Uracil, U
Thymine, T
DNA  C,T,A,G
No U
Adenine, A
RNA  C,U,A,G
No T
Guanine, G
•DNA is double-stranded (double helix), while RNA is single stranded
•RNA has a slightly different sugar backbone – ribose instead of
deoxyribose
•RNA has a lot of turns and kinks, more chaotic structure, but some
sections are closer to the outside than others…
RNA
DNA
Cell Composition
• 70-90% water
• Organic chemistry key to the construction of
cells is inherently linked to the properties of
water vs. organic compounds
• Consider 4 groups of monomers (a single,
repeated ‘building block’):
Polysaccharides
Lipids
Nucleic Acids
Proteins
Macromolecules
– Sugars
– Fatty Acids
– Nucleotides
– Amino Acids
Construction, Part 4
• Amino acids  monomer units of proteins
All amino acids have 2
functional groups – one
carboxylic acid group (COO-)
and one amino group (NH3)
Some amino acids have
hydrophobic ends, others
are acidic, some
hydrophilic, or ionizable
Bonds between the C and N
form a peptide bond, which
helps form proteins
Proteins – ‘key and lock’ concept
Peptidoglycan (aka Murein)
• Polymer consisting of both sugars and
amino acids
• Rigid material and serves a structural role
in cell wall
Cell Construction
• OK – using the building blocks we have
described, let’s make a microbe…
Predominant chemical
composition
Protein
Flagella,Pili
Function(s)
Swimming movement
Sex pilus
Mediates DNA transfer during conjugation
Protein
Common pili or
fimbriae
Attachment to surfaces; protection against phagotrophic
engulfment
Protein
Capsules
(includes
"slime
layers" and
glycocalyx)
Attachment to surfaces; protection against phagocytic
engulfment, occasionally killing or digestion; reserve of
nutrients or protection against desiccation
Usually polysaccharide;
occasionally polypeptide
Gram-positive
bacteria
Prevents osmotic lysis of cell protoplast and confers rigidity
and shape on cells
Peptidoglycan (murein) complexed
with teichoic acids
Gram-negative
bacteria
Peptidoglycan prevents osmotic lysis and confers rigidity
and shape; outer membrane is permeability barrier;
associated LPS and proteins have various functions
Peptidoglycan (murein) surrounded
by phospholipid proteinlipopolysaccharide "outer
membrane"
Plasma
membrane
Permeability barrier; transport of solutes; energy generation;
location of numerous enzyme systems
Phospholipid and protein
Ribosomes
Sites of translation (protein synthesis)
RNA and protein
Inclusions
Often reserves of nutrients; additional specialized functions
Highly variable; carbohydrate, lipid,
protein or inorganic
Chromosome
Genetic material of cell
DNA
Plasmid
Extrachromosomal genetic material
DNA
Cell wall
Prokaryote Structure
Cell wall
Nuclear material
membrane
Membrane is critical part of how food and waste are transported
- Selectively permeable
Phospholipid layer
Transport proteins
Cell Membranes
• The membrane separates the internal part of the cell from
the external  that these environments remain separate,
but under CONTROLLED contact is a key to life
Membrane Components:
•Phospholipid bilayer
•Hopanoids, which provide
additional structural stability
(similar to sterols (cholesterols)
which provide rigidity to
eukaryote cells)
•Proteins – direct transport
between outside and inside the
cell
~ 40% lipid, 60% protein
Eubacteria vs. Archaebacteria
Archaeal cell structure
Bacterial cell structure
Difference??
Let’s look more closely at the membrane, though only 8 nm thick,
it is the principle difference between these 2 groups of microbes
Archaea vs bacteria membranes
• Principle difference between these two is
the membrane
• In archaea, lipids are unique  they have
ether linkages instead of ester linkages
Membrane function
• SELECTIVELY PERMEABLE
– Passive diffusion  Gases (O2, N2, CO2, ethanol, H2O
freely diffuse through layer
– Osmosis  because solute concentration inside the cell
are generally higher (10 mM inside the cell), water
activity is lower inside, H2O comes in – increased water
results in turgor pressure (~75psi)
– Protein-mediated transport  selective and directional
transport across the membrane by uniporters and
channel proteins, these facilitate diffusion – still
following a gradient and does not require an energy
expenditure from the cell
Membrane function 2
• Active transport  proteins that function to move
solutes against a gradient, this requires energy
• Uniport, Symport, and Antiport proteins guide
directional transport of ions/molecules across
membrane – different versions can be quite
selective (single substance or class of substances)
as to what they carry
Membrane and metabolism
• As the membrane is the focus of gradients, this is where
electron transport reactions occur which serve to power the
cell in different ways
• Many enzymes important to metabolic activity are
membrane bound
H+ gradients across the membrane
• Proton Motive Force (PMF) is what drives
ATP production in the cell
Figure 5.21
Membrane functions (other)
• In addition to directing ion/molecule transport and
providing the locus for energy production,
membranes are also involved in:
–
–
–
–
–
Phospholipid & protein synthesis for membrane
Nucleoid division in replication
Base for flagella
Waste removal
Endospore formation
• Though very small, the membrane is critical to
cell function  Lysis involves the rupture of this
membrane and spells certain death for the
organism
Cell Wall
• Cell wall structure is also chemically quite
different between bacteria and archaea
• Almost all microbes have a cell wall –
mycoplasma bacteria do not
• Bacteria have peptidoglycan, archaea use
proteins or pseudomurein
• The cell wall serves to provide additional
rigidity to the cell in order to help withstand
the turgor pressure developed through
osmosis and define the cell shape as well as
being part of the defense mechanisms
• Cell wall structure
• Two distinct groups of bacteria with very different
cell walls
– Gram negative has an outer lipid membrane (different
from the inner, or plasma membrane)
– Gram positive lacks the outer membrane but has a
thicker peptidogycan layer
Peptidoglycan layer
• This layer is responsible for the rigidity of the cell wall,
composed of N-Acetylglucosamine (NAG) and Nacetylmuramic (NAM) acids and a small group of amino
acids.
• Glysine chains held together with peptide bonds between
amino acids to form a sheet
Outer membrane – Gram (-)
• Lipid bilayer ~7 nm thick made of phospholipids,
lipopolysaccharides, and proteins
• LPS (lipopolysaccharides) can get thick and is
generally a part that is specifically toxic (aka an
endotoxin)
• LPS layers are of potential enviornmental
importance as a locus of chelators and electron
shuttles
• Porins are proteins that are basically soluble to
ions and molecules, making the outer layer
effectively more porous than the inner
membrane, though they can act as a sort of
sieve
External features
• Glycocalyx (aka capsule – tightly bound
and adhering to cell wall, or slime layer –
more unorganized and loosely bound) –
helps bacteria adhere to surfaces as well
as provides defense against viruses
• Flagella – ‘tail’ that allows movement by
rotating and acting as a propeller
• Pili – thin protein tubes for adhesion
(colonization) and adhering to surfaces
Inside the cell
• Cytoplasm – everything inside the membrane
• Nucleoid/Chromosome – DNA of the organism – it
is not contained by a nuclear membrane (as
eukaryote cell)
• Ribosomes – made of ribosomal RNA and protein
 these are responsible for making proteins
• Vacuoles or vesicles – spaces in the cytoplasm that
can store solids or gases
• Mesosomes/Organelles –a membrane system
internal to the cell which facilitates protein function;
there are these structures specifically for
photosynthesis
Cell structure
Nucleoid
• Single strand of DNA, usually circular, usually
looks like a big ball of messed up twine…
• Size – smallest organism yet discovered
(Nanoarchaeum equitans) 490,889 base pairs; e.
coli 4.7 Mbp, most prokaryotes 1-6 million base
pairs (1-6 MBp); Humans 3300 MBp
• DNA is around 1000 mm long in bacteria, while the
organism is on the order of 1 mm long – special
enzymes called gyrases help coil it into a compact
form
Ribosomes
• Ribosomal RNA is single stranded
• RNA is a single stranded nucleic acid
– mRNA- messanger RNA – copies information from
DNA and carries it to the ribosomes
– tRNA – transfer RNA – transfers specific amino acids to
the ribosomes
– rRNA – ribosomal RNA – with proteins, assembles
ribosomal subunits
DNA is transcribed to produce mRNA
mRNA then translated into proteins.
RNA and protein construction
• The nucleotide base sequence of mRNA is encoded
from DNA and transmits sequences of bases used to
determine the amino acid sequence of the protein.
• mRNA (“Messenger RNA”) associates with the ribosome
(mRNA and protein portion).
• RNA (“Transfer RNA”) also required
• Codons are 3 base mRNA segments that specify a
certain amino acid.
• Most amino acids are coded for by more than one
codon: degenerage genetic code.
• Translation ends when ribosome reached “stop codon”
on mRNA.
Transcription
RNA polymeraze takes the DNA and temporarily unwinds it, templates the
transfer RNA from that, using ribonucleoside triphosphates to assemble…
Translation
• mRNA is coded for one or more specific
amino acids and moves to the ribosome to
assemble amino acids into proteins
• On mRNA, codons are 3 bases, coded to
specific amino acids
• On tRNA, the anticodon
latches to the codon
on the mRNA
Protein Formation
•
The ‘code’ on
mRNA
determines the
sequence of
protein assembly
rRNA
• Ribosomes are made of proteins and rRNA, the
tRNA and mRNA come to it andassemble the
proteins
• rRNA plays a structural role, serving as a
support for protein construction, and a functional
role
• rRNA consists of two subunits, one 30S in size
(16S rRNA and 21 different proteins), one 50S in
size (5S and 23S rRNA and 34 different
proteins). The smaller subunit has a binding site
for the mRNA. The larger subunit has two
binding sites for tRNA.
Cytoplasmic
inclusions
Where found
Composition
Function
glycogen
many bacteria e.g. E. coli
polyglucose
reserve carbon and energy source
polybetahydroxy
utyric acid
(PHB)
many bacteria e.g. Pseudomonas
polymerized hydroxy
butyrate
reserve carbon and energy source
polyphosphate
(volutin
granules)
many bacteria e.g. Corynebacterium
linear or cyclical
polymers of PO4
reserve phosphate; possibly a reserve of high
energy phosphate
sulfur globules
phototrophic purple and green sulfur
bacteria and lithotrophic colorless
sulfur bacteria
elemental sulfur
reserve of electrons (reducing source) in
phototrophs; reserve energy source in
lithotrophs
gas vesicles
aquatic bacteria especially cyanobacteria
protein hulls or shells
inflated with gases
buoyancy (floatation) in the vertical water
column
parasporal
crystals
endospore-forming bacilli (genus
Bacillus)
protein
unknown but toxic to certain insects
magnetosomes
certain aquatic bacteria
magnetite (iron oxide)
Fe3O4
orienting and migrating along geo- magnetic
field lines
carboxysomes
many autotrophic bacteria
enzymes for autotrophic
CO2 fixation
site of CO2 fixation
phycobilisomes
cyanobacteria
phycobiliproteins
light-harvesting pigments
chlorosomes
Green bacteria
lipid and protein and
bacteriochlorophyll
light-harvesting pigments and antennae