Notesale: Turn your study into money

Document Preview

Extracts from the notes are below, to see the PDF you'll receive please use the links above

---

BIOL211 BACKGROUND READING NOTES FOR LECTURES 
Physiology 1: Microbial growth and NutritionWessner et al
...
1 (pp4-6)





Microorganisms- microscopic forms of life- usually consist of a single cell and include
bacteria, archaeons, fungi, protozoa, and algae
...
Can cause infectious diseases
...

Good and harmful bacteria ->

 beneficial microbes associated with our bodies help us digest food and protect us from
potentially harmful microbial invaders
 harmful bacteria, viruses, fungi and protozoa kill millions and sicken billions
some microbes cause crops to fail/  whilst some provide nitrogen to plant roots through
symbiotic relationships
some microbes cause food to rot/  whilst others carry out fermentation









The Basis of Lifeo Metabolism- A controlled set of chemical reactions that extract energy and nutrients
from the environment, and transform them into new biological materials
o Growth- An increase in the mass of biological material
o Reproduction- The production of new copies of the organism
To accomplish these tasks, organisms contain a biological instruction set to guide their
actions
...
Other
features that living organisms share include:o Genetic variation- allowing the possibility of evolution or inherited change within a
population, through natural selection over the course of multiple generations
o Response to external stimuli and adaptation to the local environment (within genetic
and physiological constraints)
o Homeostasis= active regulation of their internal environment to maintain relative
constancy
This list doesn’t always represent a complete description of what it means to be alive! For
example, in endospores (can remain dormant for years)
...
A free-living unicellular,
or single-celled, organisms can carry out all the necessary functions of metabolism, growth,
and reproduction without physical connection to any other cells
...
g
...
During periods of nutrient
depletion, however, individual cells aggregate and form a complex structure, with cells
differentiating to assume specialised tasks
...
Chapter 6 (pp165-201)  covers the first four lectures!
INTRODUCTION



Microorganisms are capable of reproducing faster than any of the larger forms of life on earth,
and microbial growth has enormous consequences for the biosphere and human health
Microbes play integral roles in biogeochemical cycling, transferring important elements like
carbon and nitrogen from one molecular state to another
...

Cells also require- potassium, sodium, magnesium, calcium and iron (are the most abundant
cations in microbial cells)
...
K+, Na+, Cl- have important functions in controlling osmotic
balance, and in charge dependent with macromolecules
Mg2+ associates with ATP, and is a key component of many enzymes and protein-nucleic
acid complexes, such as the ribosome
...
Iron is frequently limiting for
growth in natural habitats, where soluble iron can be in short supply
...

Micronutrients- zinc, cobalt, molybdenum, copper and manganese are also needed for
microbial growth
...
Most microbes need so little of these metals that they often are not
intentionally added to culture media

Acquisition of nutrients








Acquiring carbon and incorporating in into macromolecules is critical for a growing cell – the
process by which a cell imports a molecule and incorporate it into cellular constituents is
called assimilation
...
They convert the carbon present in
molecules such as carbon dioxide into organic molecules, this is called carbon fixation
...

Heterotrophs- can’t assimilate carbon from inorganic carbon molecules
...
Heterotrophic microbes have evolved to acquire carbon from
many kinds of molecules- carbohydrates, amino acids, lipids, organic acids, alcohols and
more
...
Certain microbes can convert the dinitrogen gas (N2) present in the
atmosphere into ammonia (NH3), through a process known as nitrogen in the form of
ammonia (NH3)
...
Once a cell obtains ammonia through any of these avenues, it can
incorporate the nitrogen into the amino acids glutamate and glutamine
...


Acquisition of energy






Cells need energy to drive their metabolism
Microorganisms require energy in various ways
...
Many phototrophs, use the energy
generated from p/s for carbon fixation, incorporating carbon dioxide into biological molecules
such as carbohydrates
...

Chemotrophs- acquire energy through the oxidation of reduced organic or inorganic
compounds they have acquired from the environment- in many cases where organic
molecules are used as a source of energy, the same molecule serves both as a carbona
energy source
...
g
...


Electron source





All microorganisms require a source of electrons
...

Organotrophs- acquire electrons from organic molecules such as glucose
Lithotrophs- remove electrons from inorganic reduced molecules, such as Ferrous Iro,
elemental sulfur, hydrogen gas, hydrogen sulphide, ammonium and nitrite
...


FACTORS AFFECTING MICROBIAL GROWTH






Microbial growth rate and yield are determined by the interaction of environmental factorsboth physical and chemical and the genetically encoded traits of an organism
...

While doing this they must extract usable energy from their surroundings in order to drive
chemical reactions
...

Temperature, pressure and illumination can also affect cell structure and function and thereby
influence growth rates

Effects of nutritional factors on microbial growth








When a molecule enters the cytoplasm of a cell, it may contribute to growth if it’s used to
synthesise cellular components or it may contribute to the production of ATP
...

The rate at which the cell can acquire nutrients, process them into macromolecules, and then
assemble them determines the rate of cell growth
Before the cell can divide it must:
o (1) fully duplicate its genome
o (2) double its mass approximately (largely composed of proteins, nucleic acids, lipids
and cell wall
...
or cellular machinery or cell size will steadily decrease every generation
...


Nutrient Type










The type of nutrients a microbe accesses affect its growth rate and yield
Prototrophs- synthesise all necessary cellular constituents from a single organic carbon
source and inorganic precursors
...
Instead certain organic precursors
are acquired from the environment  as a result the growth of these organisms is limited to
niches in which appropriate organic molecules can be found
...
This is because synthesis of macromolecular
precursors such as amino acids “from scratch” using nitrogen from ammonia and carbon
derived from glucose is costly in terms of ATP
Cells that import amino acids can devote their energy to simply linking them together into
proteins
A cell that can’t import amino acids, in contrast, must use a great deal of energy, carbon and
nitrogen to synthesise them net result= less protein assembled per unit of energy expended
and less cellular material produced per unit time
...

If the concentration of a critical nutrient is low, then one or more biochemical pathways
may operate at less than their maximal potential
Once the bacteria are dividing as fast as they possibly can, further increases in nutrient
concentration have no effect on growth rate

Effects of oxygen on microbial growth













Aerobes- require oxygen for growth
...
Growth rates for most obligate
aerobes are maximal or at near atmospheric O2 concentrations, and decline as the
oxygen conc
...

Anaerobic growth- occurs without the use of oxygen
Aero-tolerant anaerobes- don’t use oxygen for respiration but are not harmed by it
Obligate anaerobes- can’t grow in the presence of oxygen / lack adequate defences
...

Aerobic microbes depend on multiple defences against oxidative damage since reduction of
oxygen during respiration generates some of these potentially dangerous by-products,
undergo aerobic respiration need defences against toxic oxygen species generated by
biochemical and abiotic reactions
...

Microaerophiles- show maximal growth rates at oxygen concentrations lower than that
typically in the atmosphere  these microbes are adapted to relatively stable zones where
oxygen is depleted by microbial growth and diffusion of atmospheric oxygen is restricted
...


Effects of pH on microbial growth











pH  expresses the hydrogen ion activity or concentration of H+ ions of a solution
pH scale is an inverse logarithmic function (pH=-log[H+]) ranging from 0 to 14
...

Microbial growth rates can be affected significantly by pH
...

Neutrophiles- grow optimally in environments with pH close to neutrality (5
...
5)
Acidophiles- prefer significantly acidic habitats (pH <5
...
 Aerotolerant anaerobes generate energy through fermentation often producing lactic acid
as a primary fermentation product
...

Because their fermentative metabolism is so inefficient, they grow best in high sugar
concentrations
...
These bacteria are often found in
anaerobic habitats rich in sugars and other organic materials
...
5)
Microbes can also play an active role in altering the pH of their environment
...

It is common for microbes to maintain higher solute concentrations in their cytoplasm than in
the surrounding environment, so some osmotic pressure is normal
...

For microorganisms to grow, water must not only be present, it must also be accessible
...

Most bacteria require an aw of greater than 0
...


Effects of temperature on microbial growth














Mesophiles- grow at temperatures considered “normal”- between 10C and 40C
Psychrophiles- grow at temperatures below 10C
Thermophiles/hyperthermophiles- grow optimally at temperatures above 55C-80C
...

o Increasing temperature raises the thermal energy of molecules, making them more
likely to react
o Decreasing temperatures lowers the available energy
At lower temperatures proteins fold more slowly and dynamic changes in structure that may
be necessary for proper function are restricted
At higher temperatures the effects of thermal energy may overcome some of the weaker
chemical interactions that help to stabilise protein structure
Fluidity of the membrane bilayer is also affected by temperature
...
The progressive “freezing” of a
membrane can negatively affect critical transmembrane processes, such as nutrient
transport, electron transport and maintenance of electrochemical gradients
...

Because the effects of both low and high temperatures on membranes are somewhat
dependent on lipid composition, many microbes actively adjust lipid composition in response
to temperature
...


GROWING MICROORGANISMS IN THE LABORATORYMedia for microbial growth





Liquid and solid forms of growth media, combinations of nutrients designed to allow the
growth of microorganisms, may be used
...
5%, is mixed with water and
other media components
...
The high temperature also destroys most, if not all microbes present
in the solution, thereby sterilising it
...
The molten agar will transition to a solid gel when the medium cools
to around 40C
...

In the lab, pathogens often prefer, or even require, a rich medium with abundant sugars,
amino acids and vitamins
...

o Other common components of complex media include yeast extract composed of
beef muscle tissue that has been soaked and boiled and the soluble components
extracted
...


Specialised media









Selective media- designed to allow the growth of only certain target organisms
...
g
...
Both of these species are
require very nutrient rich media, but grow so slowly that other bacteria and
fungi easily overwhelm them
...

o Selective mediums can encourage the growth of particular bacteria’s and discourage
the growth of others that aren’t needed
Differential media- allows a researcher to distinguish two, often related, types of bacteria
o MacConkey agar- the selective agar also allows some important species to be
differentiated by colony colour
...
--> hemolysis causes a clear
or greenish zone around the bacterial colony
Enrichment media- contains nutrients or other components designed to favour the growth of
particular microbes
...

Rich media enrich for microbes that are nutritionally versatile, have the fastest and/or highest
capacity transport systems and are adapted to high-nutrient fluxes
...

o Incubating plates aerobically in a standard atmosphere of 20% oxygen selects
against anaerobes and microaerophiles, while incubating anaerobically does the
opposite
...

Advantages of a solid medium are that cells are held in place
Isolated colonies should contain a single type of microorganism; all cells within the colony
should be descended from a single cell




Cultivation- independent methods based on nucleic acid detection and metagenomics can
allow identification and characterisation of uncultivated microbes
Members f the microbial consortia can evolve obligate dependencies with other members of
the community that preclude their isolation as individual species

Measuring and monitoring microbial populations










In the lab- microbial growth can be controlled and monitored
The actual number of cells in a sample can be determined by a direct count, Petroff-Hausser
counting chamber
The number of viable cells can be determined by adding bacteria to an agar medium, either
via the spread plate or pour plate method
...
As the population density increases, the absorbance, or optical density increases
Microbial populations growing in a batch culture tend to follow a growth curve including 4
phases:o Lag
o Log/ Exponential
o Stationary
o Death
By analysing the growth kinetics, the generation time, growth rate, and growth yield of the
microorganisms can be calculated
Continuous culture operate as open systems
...
In the lab- this destruction occurs in an autoclave
...

Antiseptics can be used on skin
Antimicrobial compounds can be classified by structure or mechanism of action
...


Physiology II: Batch Culture
Wessner et al
...
Chapter 6 (pp165-201)  covers the first four lectures!

Physiology IV: Example data analyses
Wessner et al
...
pages 406-415 
CENTRAL PROCESSES IN ATP SYNTHESISHow do cells make ATP?





ATP= the primary currency used to power virtually every activity of cells from bacteria to
human
...

This reaction can occur because the transfer of the phosphate group from a molecule with
high potential free energy (the intermediate) creates one of the lower potential free energy
(ATP)
...

A commonly used pathway for the production of ATP is the catabolism of glucose in
glycolysis
...

o One of the intermediate molecules generated during glycolysis is 1-3
biphosphoglycerate
...

o The catabolism of 1-3 bipshophoglycerate releases an inorganic phosphate (Pi) and
produces 3-phosphoglycerate  the energy released is used to produce a covalent
bond between Pi and ADP to produce ATP
...

Oxidative Phosphorylation- uses the chemiosmotic process and requires the transfer of
electrons through the electron transport system, which generates a proton gradient
...

o Used by chemotrophs
Photophosphorylation- also produces ATP using a proton gradient and an electron transport
system
...

o Chemotrophs- acquire energy through oxidation of preformed chemicals that they
obtain from their environment
o Phototrophs- acquire energy by capturing photons of light to raise electrons to higher
energy levels that can drive otherwise unattainable redox reactions
o Organotrophs- remove electrons from organic molecules, such as glucose, whereas
lithotrophs remove electrons from inorganic reduced molecules such as H 2S, H2 or
elemental sulfur
Heterotrophs obtain carbon from organic molecules, such as sugars, obtained from their
environment
Autotrophs use inorganic carbon compounds such as CO 2

CARBON UTILISATION IN MICROORGANISMS
How do microbes utilise organic carbon?




Carbo= most abundant element in biological macromolecules, acquiring it is critical for cell
growth and replication
Assimilation= the general process by which cells import a molecule and incorporate it into
cellular constituents
Heterotrophic microbes have adapted to transport and incorporate carbon from many kinds of
organic molecules – carbohydrates, amino acids, lipids, organic acids, alcohols and more
...
Another ATP molecule is invested after
isomerization to fructose 6-phosphate, to produce a high-energy molecule of
fructose 1, 6- bisphosphate, which is cleaved into two three carbon
compounds
...

2
...

4
...

6
...

8
...

10
...
Catabolism of the two glyceraldehyde 3-phosphate molecules will
produce even more potential free energy; four ATP molecules are
synthesised by substrate-level phosphorylation
...
Before any energy earning can be made, energy must
be invested
 PHASE II- generates energy in the form of electrons as well as ATP, and
begins with the first oxidation
...
NADH is produced From NAD+
Substrate- level phosphorylation produces ATP and releases 3-phosphoglycerate
3- phosphoglycerate is reconfigured
2- phosphoglycerate is dehydrated, forming the high energy molecule phosphoenolpyruvate
Substrate-level phosphorylation produces ATP and releases pyruvate

The Entner- Doudoroff Pathway
o Less commonly used for glucose catabolism but is often used for catabolism of other
carbohydrates containing aldehyde groups, such as gluconate, which cannot be
readily processed by the EMP pathway
...
A phosphate group is added from ATP to form glucose 6-phosphate
2
...
NADPH is
produced from NADP+
3
...
2- keto3-deoxy-6-phospholuconate
4
...
Pyruvate
is produced and glyceraldehyde 3-phosphate is further converted
5
...
Glucose 6- phosphate is oxidised to 6phosphogluconate and generates NADPH in the process
...
The ribulose backbone is converted to two
different pentoses, ribose and xylulose
...
Erythrose phosphate can be combined with
xylulose phosphate to generate fructose 6-phosphate and glyceraldehyde 3phosphate
...
The result is the conversion of three molecules of glucose, to two
molecules of fructose 6-phosphate and one glyceraldehyde 3-phosphate, with the
production of six molecules of NADPH
...
The fate of electrons carried by NADH or other reduced
electron carriers generated from glycolytic pathways depends on the genetic capacity
of the organism and the conditions it experiences
Fermentation
o Many organisms such as lactic acid bacteria (LAB) are capable of using fermentation
under low-oxygen conditions
...
Fermentation regenerates NAD+, allowing glycolysis to
continue to produce ATP
o Produces only two moles of ATP for one mole of glucose
o To keep up with energy needs, glycolysis occurs at a much faster rate during
fermentation than during respiration
...
, pp
...

o The fate of CH4 is also important to consider because of its high-infrared absorbance
...
These microbes are a subgroup of the
methylotrophs, members of the Proteobacteria that can oxidise organic compounds
like methane, formate, and methanol that do not contain C-C bonds
...

o The methanotrophs metabolise methane, converting the carbon in this molecule into
forms that can be utilised by other organisms
...

Acetotrophic methanogens produce methane from acetate (CH3COO-)
...

o CH4 + SO42-  HC03- + HS- + H2O

Physiology VI: Photoautotrophy
Wessner et al
...


Wessner et al
...

o First step- photophosphorylation
o Second step- uses the ATP produced by photophosphorylation to fix carbon dioxide
to make carbohydrates
o Some photoorganoheterotrophs can carry out photophosphorylation to make ATP,
but do not use it for carbon fixation, instead obtaining carbon from organic sources
o Chlorophyll based photosynthesis is a bacterial invention that has found its way into
eukaryal algae and plants thanks to endosymbiotic cyanobacteria that evolved into
chloroplasts
Phototrophy- found in five distinct lineages within the domain Bacteria
...
The only Gram-positive group is
the genus Heliobacterium, belonging to the phylum Firmicutes
...
Other bacteria do not
generate O2 from photosynthesis

Photophosphorylation: ATP synthesis and the “light” reactions







Photophosphorylation- converts light energy to chemical energy that can be used by the cell
 reactions are often referred to as light reactions as they require the presence of light
Bacteria do not contain chloroplasts (where the light reactions take place)
...
In Gram negative
bacteria, the lumen is an extension of the periplasm
Photophosphorylation depends on light-absorbing photo-pigments to capture photons
...

Anoxygenic Photosynthesis- Photosystem I is the sole photosystem in anaerobic
photosynthetic heliobacteria and green sulfur bacteria and is responsiblefor anoxygenic
photosynthesis
o Chlorobium tepidum: gram negative, carries out carbon fixation via a reductive TCA
cycle and anoxygenic photosynthesis using sulphides such as H 2S, as electron
donors






Oxygenic Photosynthesis- Cyanobacteria and chloroplasts combine PS I with PS II in
oxygenic photosynthesis, a non-cyclic process that obtains electrons from water and
produces both reducing power and proton motive force
...

2H2O  4H+ + 4e- + O2
The structure and function of PS II in oxygenic photosynthesis differs from Rhodospirrilum in
that, instead of the excited electrons ultimately flowing back to the PS II in a cyclic pathway,
they are passed to PS I
...
This
product is then converted into a variety of macromolecules, such as hexose sugars
and amino acids
Ribulose bisphosphate carboxylase  Rubisco
...
This is the actual fixation step and is
referred to as the carboxylation phase of the Calvin cycle
The enzyme Rubisco adds a carbon (carboxylation) to ribulose 1,5-bispohsphate and cleaves
it, producing 2 molecules of 3-phosphoglycerate for each ribulose 1,5- bisphosphate
3-phosphoglycerate is phosphorylated to produce 1,3- bisphosphoglycerate, which is reduced
to glyceraldehyde 3-phosphate
...
This begins the regeneration phase
...
Some organisms can run the cycle in reverse to consume CO 2 to produce carbon
compounds at the expense of ATP and electrons, usually in the form of NADH
...

Intermediates of the reductive cycle can also be used for biosynthesis
The reductive TCA cycle goes in the opposite direction to the TCA cycle starting with
oxaloacetate
...
A
product is the two-carbon compound acetyl- CoA, which enter into further biosynthetic
reactions, depending on the metabolic capabilities of the organism
...
Rev
...
g
...
g
...
g
...

Aerobic and anaerobic (often denitrifying) chemolithoautotrophic bacteria and archaea use
the reducing equivalents for both respiration and CO2 fixation, while anaerobic
photolithoautotrophic bacteria use reductants mainly for CO 2 fixation
...


Extreme Environments I:Thermo/ psychrophiles
Wessner et al
...

Most of the cultures crenarchaeotes fall into one of these two categories

THE THERMOPHILES AND HYPERTHERMOPHILES








Most thermophilic and hyperthermophilic crenarchaeotes have been isolated from thermal
springs and geysers
...
Many of these hyperthermophiles also are acidophiles, growing in low pH
environments
...
 isolated from a volcanic crater near Naples, Italy, this microbe has
an optimal growth temperature of approximately 80ᵒC/ additionally it grows optimally at pH
3
...

Other members of this phylum have been isolated from deep sea hydrothermal vents
...

Life at high temp
...
Plasma membranes containing tetra-ether
lipids or lipid monolayers may provide extra stability at high temps
...

Elevated temperature also generally causes the denaturation of proteins
...
It appears
that hyperthermophiles may employ two distinct mechanisms to maintain protein integrity
...
The increased ἀ-helical content and increased
proportion of arginine and tyrosine may lead to strengthened interactions between
amino acids  allowing the protein to maintain its shape at elevated temperatures
Hyperthermophiles also rely on a series of molecular chaperones- proteins that help fold
proteins or refold denatured proteins, to maintain the functionality of their proteins
...
Some species possess thermostable DNA-binding proteins that
increase the melting temperature of a double-stranded DNA
...
By increasing the degree of supercoiling,
these enzymes greatly increase the temperature at which the DNA unwinds and
denatures

THE MESOPHILES AND PSYCHROPHILES




These organisms grow optimally between 15ᴼC and 40ᴼC or at temps
...
Since
the early 1990’s, researchers have detected crenarchaoter ribosomal RNA sequences in
various temperature and cold marine environments
Several pieces of evidence suggest that these organisms are major contributors to carbon
cycling in the oceans
o Nitrosopumilus maritimus (marine crenarchaeote) this organism may contribute to
nitrogen cycling
 N
...
Genome sequencing studies shows that it contains an ammonia
monooxygenase gene, the product of which converts ammonia to
hydroxylamine, the first step in the nitrification process
...
These organisms produce
methane from a variety of substrates, such as carbon dioxide or various simple
organic molecules like methanol (CH3OH) and formate (CHOO-)
...
These organisms, then, are autotrophs,
incorporating inorganic carbon into organic molecules
...
g
...
All identified methanogens are strict
anaerobes, organisms that can survive only in anoxic, or oxygen – free
environments
...




o

The large intestine contains a great diversity of microbes, including the
methanogen Methanobrevibacter smithii
...

 Anoxic habitats in which methanogens exist can be fairly diverse
...
Some methanogen species like
Methanothermus fervidus are rods, others like Methanculleus olentangii are
cocci
...
5M
...
Because these lakes have no outlets, water
only leaves through evaporation
...

 Additionally, temperatures are very low
...
The high intracellular potassium levels supply the cell
with a type of osmotic balance, thereby preventing the efflux of water
 But high intracellular levels of potassium cause problems:1
...
The increased
potassium levels would disrupt the hydrogen bonds that hold
together the two strands of a double-stranded DNA molecule
2
...
Again proteins tend to denature when
placed in high salt environments
How do Halophiles avoid this problem?  First, the genome has a high G-C
content
...
Secondly, the proteins are highly acidic,
containing a large number of aspartic acid and glutamic acid residues
...



Members of this genus also have a special way of obtaining energy
...
salinarum obtain energy via phototrophy, the
acquisition of energy from sunlight
...
Absorption of light by
retinal results in the transfer of an electron by bacteriorhodopsin
directly into the periplasm
...
These protons then flow back into the
cell through a typical ATP synthase H+ channel, generating ATP
...
7
...


Wessner et al
...

Temperature affects the rate of chemical reactions
...
Decreasing temperature lowers the available energy,
so we could expect growth to slow as temperature declines below a microbe’s optimal growth
temperature, as biochemical reactions slow
...
At lower temperatures, proteins fold more slowly, and dynamic changes in structure
that may be necessary for proper function are restricted
...

Fluidity of the membrane bilayer also is affected by temperature
...

o The progressive “freezing” of a membrane can negatively affect critical transmembrane
processes, such as nutrient transport, electron transport, and maintenance of
electrochemical gradients
...
The organisation and structural integrity of a membrane bilayer,
including its constituent proteins, also can break down at higher temperatures
...


Trivedi et al
...
J
...
Mol
...
4: 61-69
...
These include high CG content in the coding sequences, nucleotide arrangement of
purine-purine and pyrimidine – pyrimidine, methylation of nucleotides, histone/ histone like
proteins, reverse gyrase, cations, etc
...
Strategies are
adopted at the level of DNA are naturally reflected in RNA in thermophiles
...
All these factors may differ from taxa to taxa as no single or all the factors
together can be universally attributed for providing thermal stability to nucleic acids
...
(2006) Gen
...
Res
...

 Among several important contributing factors for stability of proteins are CG-rich codons, the ration
of charged amino acids compared to uncharged amino acids, ionic interactions, amino acid
preferences and their distribution, post-translational modifications, and solute accumulation
...
This is exemplified in the
case of differences in strategies adopted by soluble proteins and membrane proteins
...


Feller (2003) Cell Mol
...
Sci
...









Psychrophiles are cold-loving organisms successfully colonize cold environments of the Earth’s
biosphere
...

Emerging evidence suggests that psychrophilic enzymes utilise an improved flexibility of the
structures involved in the catalytic cycle, whereas other protein regions if not implicated in catalysis
may or may not be subjected to genetic drift
...
e
...

The flexible structure of psychrophilic enzymes can provide enhanced ability to undergo discrete
and fast conformational changes at low temperatures imposed by the catalytic events
...


The pH scale is an inverse logarithmic function (pH= - log[H+]) ranging from 0 to 14
...
5-8
...
5)

Alkalophiles prefer alkaline habitats (pH>8
...
These aerotolerant anaerobes generate energy through fermentation, often producing
lactic acid as a primary fermentation product
...

Because their fermentative metabolism is so inefficient, they grow best in environments with high-sugar
concentrations
...
These bacteria are often found in anaerobic habitats

Microbes also play an active role in altering the pH of their environment
...
15: 165-171
...

 Some acidophiles are involved in the catalysis of sulphide mineral dissolution, resulting in high
concentrations of metals in solution
 Acidophiles have a variety of intrinsic and active metal resistance systems that likely combine
to permit their growth in very high metal concentrations

Horikoshi K (1999) Microbiol
...
Biol
...
63: 735-750
 Alkaliphiles grow optimally or very well at pH values above 9 but can’t grow or grow only slowly
at the near neutral pH value of 6
...

 They can be isolated from normal environments such as garden soil, although viable counts of
alkaliphiles are higher in samples from alkaline environments
 The cell surface may play a key role in keeping the intracellular pH value in the range between
7 and 8
...
Appl
...
110: 851-861
...
, Deinococcus radiodurans, Rubrobacter radtiolerans and Thermococcus
gammatolerans are all examples of radioresistant microorganisms with the ability to survive
and grow under high doses of radiation
...

 The application of radioresistant microorganisms to manufacturing and service industries and
their use to produce products beneficial to man: using the identification and study of gene
sequences in its DNA, the study of proteins expressed by genes within the organism, with
applications in the understanding of disease and in drug development and the measurement
of the metabolites of low molecular weight in the organism’s cells at a specific time under
specific environmental conditions is being highlighted
...
Microbiol
...

 In classical models of radiation toxicity, DNA is the molecule that is most affected by ionizing
radiation (IR)
...
This would allow an irradiated cell to protect sufficient enzymatic
activity needed to repair DNA and survive
...
Rev
...

 Deinococcus radiodurans and other radiation resistant bacteria accumulate exceptionally high
intracellular manganese and low iron levels
...
radiodurans might erve as
antioxidants that reinforce enzymic systems which defend against oxidative stress during
recovery

Extreme Environments IV: Storage states
Wessner et al
...
The endospores are largely metabolically inert structures that exhibit increased
resistance to many harsh environmental conditions e
...
desiccation, UV light exposure and
high temperatures
...

Endospores shut down their metabolism completely and compact chromosomal DNA tightly
with protective proteins
...
Microbiol
...





Spore formation in bacteria poses a number of biological problems of fundamental
significance
...

Sporulation is one of the best understood examples of cellular development and
differentiation
...


Knaysi (1948) Bacteriol
...
12: 19-77
...


HETEROCYSTS:
Wolk et al
...
Rev
...

Heterocyst structure and metabolic activity function together to accommodate the oxygensensitive process of nitrogen fixation
Heterocyst- forming cyanobacteria differentiate highly specialised cells to provide fixed
nitrogen to the vegetative cells in a filament
Heterocysts are typically distinguishable from vegetative cells by their somewhat larger and
rounder shape, diminished pigmentation, thicker cell envelopes, and usually prominent
cyanophycin granules at poles adjacent to vegetative cells
...

Mature heterocysts provide the microoxic environment required for nitrogen fixation, spatially
separating oxygen- evolving photosynthesis in vegetative cells from nitrogen fixation
...
(1982) J
...
149: 354-360





Exospores form at the tapered end of the vegetative cell are resistant to heat and dessication
but, unlike bacterial endospores, contain no dipicolinic acid
...

Formation of exospores by M
...
Once formed they are resistant to heat and dessication
...
Constriction of the bud resulted in a structure similar to that of
the immature exospore but attached by an edge to the vegetative cells so that the whole
structure has bilateral symmetry

Biogeochemistry I: Sulfur cycle
Wessner et al pages 406-415


As above

Wessner et al pages 470-480






The ability of aerobic methanotrophs to oxidise methane depends on methane
monooxygenase (MMO), the enzyme responsible for converting CH4 to CH3OH (methanol)
...

o In a reaction requiring a reductant, MMO adds an O atom derived from oxygen to
methane producing methanol plus water
...

o The requirement of oxygen as a reactant in the MMO reaction is the reason these
bacteria are obligate aerobes, and of course the oxygen is the terminal electron
acceptor in their energy metabolism as well
...
Additionally some
methanotrophs produce soluble MMO, or sMMO,that is not membrane associated
...
They have been isolated from muds,
swamps, peat bogs, sediments, fresh and saline waters, rice paddies, sewages sludge and
soils
Methane oxidation by the methanotrophs reduces the amount of methane that these sources
ultimately release to the atmosphere
...

o At this location they can obtain methane from anaerobic methanogenic activity and
oxygen from the atmosphere or aerated water
o Methanotrophic populations in lakes with methane-emitting sediments tend to occupy
a relatively narrow band of water where the methane rising up through the water
column meets oxygen diffusing downward from the lake surface
...

Clearly, the oxidative reaction sequence catalysed by the aerobic methanotrophs and
involving MMO cannot be at work here as no oxygen is present so another means of oxidising
the methane molecule must be involved
...

o ANME archaeons occur in close association with sulfate-reducing bacteria (SRB) like
Desulfobacterium autotrophicum
o Together the microbes accomplish AOM in anoxic zones where areas of sulfate
reduction and methanogenesis intersect
o In these areas methane released from methanogenesis encounters available sulfate
(SO42-) 
 CH4 + SO42-  HCO3- + HS- + H2O

CYCLING DRIVEN BY NITROGEN METABOLISM
How do microbes influence the cycling of nitrogen?





Nitrogen- mostly found in the atmosphere, mostly in terrestrial plant material, but also in land
animals and marine plants and animals
Within these locations major forms of nitrogen include organic nitrogen within biomass and
inorganic nitrogen such as NH4+ (ammonium), N2 (dinitrogen), NO3- (nitrate), NO2- (nitrite) and
N2O (nitrous oxide) these molecules contain nitrogen in different oxidation states – these
molecules additionally, may or may not be metabolically available to cells
The nitrogen cycle consists of several key steps – each carried out by specific organisms with
characteristic metabolic qualities
o Nitrogen fixation- atmospheric N2 is reduced to NH4+
...

 Nitrates can be directly assimilated by plant and a number of microbes, thus
becoming incorporated into biomass, or they can be converted back into
nitrogen in a process denitrification
...
Some are aerobes, whilst others are anaerobes
...

Terrestrial or aquatic
...

Ammonium oxidisers, such as members from the Betaproteobacterial Nitrosomonas
etc convert ammonium to nitrite
...
Hydroxylamine is then oxidised into nitrite by the
heme-containing enzyme hydroxylamine oxidoreductase
Nitrifiers carry out nitrite oxidation- second step of nitrification
...
In these systems the
produced nitrate is processed by denitrifiers, resulting in nitrogen returning back into
the atmosphere
Denitrification- heterotrophic process in which an organic substrate, under anoxic conditions,
is oxidised with nitrate as a terminal electron acceptor
...


HUMAN IMPACT ON THE NITROGEN CYCLE






Burning of fossil fuels and the use of synthetic fertilisers
o Haber-Bach industrial process- used for the production of synthetic nitrogen fertilisers
o This process relies on a series of chemical reactions that extract H2 from CH4 so it
can be combined with N2 under high temperature and pressure to form NH3
...
These reservoirs also
include carbon dioxide produced via respiration and decomposition of organic matter and
fixed by autotrophs
Water represents another major source of oxygen
Living and dead biomass constitute another reservoir of actively cycled oxygen atoms
...


CYCLING OF SULFUR AND PHOSPHORUS


Very little sulfur or phosphorus exists in the atmosphere- the major reserves of these
elements exist in rock or are dissolved in the oceans

o












Weathering of rocks can release phosphate ions from phosphorus-containing
minerals like apatite
...
Phosphate ions released from rocks into the soil and water can be taken up by
plants- once incorporated into organic molecules, phosphorus can be transferred between
organisms by process of ingestion and decay
The normal concentration of usable phosphorus in the soil is typically quite low- to counteract
this, farmers often apply large amounts of phosphate- containing fertilisers to their fields, to
increase the growth of their crops
Bacteria may also help aid in this process- such as Pseudomonas, Bacillus etc have the
ability to solubilise inorganic phosphate compounds  these same bacteria have been shown
to improve the growth of plants
...

Very little sulfur exists in the atmosphere – in the absence of biological cycling sulfur leaves
rocks via weathering and travels to the ocean, where it exists as SO 42-
...
The geochemical cycling of sulfur interacts with the
biochemical cycling of sulfur
...

Because of its energetic cost, the enzymatic machinery for nitrogen fixation is highly regulated
and is only expressed when alternative forms of nitrogen are exhausted or unavailable
The conversion of nitrogen gas nto compounds for cell use is termed nitrogen fixation
o It is purely a microbial process, carried out only by bacteria and archaea
...
These bacteria fix nitrogen
to supply their own and their host plant’s nitrogen needs  in return they receive
organic compounds from their host to supply their carbon and energy requirements
The key enzyme for nitrogen fixation is nitrogenase, this catalyses the reaction:

N2 + 8H+ + 8e-  2NH3 + H2







This reaction is accompanied by the hydrolysis of at least 16 ATP molecules
It is thought that nitrogen gas displaces hydrogen in the enzyme active site
Nitrogenase= consists of two types of subunits: ditrogenase reductase – this provides
dinitrogenase with reducing power and ditrogenase which actually reduces nitrogen gas
...
For each
electron transferred, two ATP molecules are hydrolysed
o The FeMo-co cofactor is extremely sensitive to oxidation – making it very difficult
to operate in an aerobic environment
...
PSI is not
degraded, but instead shifts to a cyclic electron transport pathway, like
that used by purple bacteria with PSII
...
In return,
heterocysts share nitrogen in the form of amino acids
...
These have to enter the cytoplasm through a transport system
...
There are two
common microbial pathways for incorporation of ammonium:
 The GS-GOGAT pathway
 The GDH pathway
o GS-GOGAT pathway= uses a combination of two enzymes: glutamine synthetase
(GS) and glutamate synthetase
...

o GDH pathway= uses the enzyme glutamate dehydrogenase to add ammonium to ἀketoglutarate
...


o




Both of these pathways are found in many microbes
...
GS has a higher affinity for ammonium than glutamate
dehydrogenase, so GS is more active at submillimolar NH4+ concentrations
...

GDH reaction is reversible, so the enzyme can function for catabolic deamination of
glutamate, generating NADPH and ἀ-ketoglutarate

UTILISATION OF NITRATE AND NITRITE






Nitrate can be used as a terminal electron acceptor for anaerobic respiration  “dissimilative
nitrate reduction”
o “assimilative nitrate reduction” = nitrate is reduced to ammonia
...

Production of nitrite sets the stage for nitrite reductase to go into action
...
Each transfer is accompanied by uptake of two
protons
...

o Denitrifying bacteria also generate nitrite from nitrate, but the dissimilatory nitrite
reductase instead generates nitric oxide, which is further reduced to nitrous oxide and
ultimately to dinitrogen
...
The most common inorganic
form of sulfur in most environments is sulfate ion  this is highly oxidised and must be
extensively reduced before it can be incorporated into cysteine
...

Sulfate- is a very stable molecule, to which it is difficult to add electrons, to make it a better
electron acceptor, sulfate is covalently attached to ATP by the enzyme ATP sulfurylase,
generating adenosine 5’ phosphosulfate (APS), this is converted into to phosphoadenosine
5’- phosphosulfate (PAPS)
...
The sulphite is reduced
further to hydrogen sulfide – potentially toxic intermediate that is quickly used to displace
acetate in acetylserine to generate cysteine
...
Sulfate reducers that use the dissimilatory pathway for energy metabolism can supply
other microbes with reduced sulfur, and often live in close proximity to anaerobic phototrophs
that consume hydrogen sulfide as an electron donor for p/s
...
These
elements can come from a variety of sources
...
Over time, microorganisms initially present in relatively low numbers will
proliferate in different sections of the tube
...

Oxygen levels decrease, though, as the distance from the surface increases
...

o Deeper down in the anoxic regions at the bottom of the column, anaerobic sulfur
reducing bacteria thrive
...


Wessner et al pages 474-475



As above on nitrification and denitrification
As above on human impacts on the nitrogen cycle

Wessner et al pages 480-418
As above on the cycling of sulfur and phosphorus



---