Lecture 3: Microbial Growth And Nutrition Flashcards
Macronutrients
- Elements required in LARGE amounts to build macromolecules (b/c you req. a lot of it)
- The building blocks of cell material
- screw to build house is a macronutrient b/c you need A LOT of it to build the house
List some macronutrients and how much of the dry weight of the cell do they make up?
C,H,O,N,P and S makeup >90% of the dry weight of the cell
What does “dry weight of the cell” mean?
dehydrated
- water makes up 60-80% of the weight of cell, this will skew the #’s too much so you consider cell in dehydrated state to see how the atoms will be req.
Protein
Polymer of made of building blocks – amino acids
C,H,O,N(andS)
• > 50% of cell dry weight
Why do proteins make up greater than 50% of the cells dry weight?
proteins are the work horse of the cell (enzymes, structural proteins, transmembrane proteins)
Protein–C,H,O,N(andS); which are from where?
C,H,O,N - backbone of an AA
S - 2/20 AA’s (cysteine/methionine)
Lipids
C,H,O(andP)
- phosphate; head groups of phospholipid structure
- C H- contribute to non polar character
- O; within carboxyl group and glycerol component of neck of structure
- Building blocks = fatty acids and glycerol
- Ex) Phospholipids
Carbohydrates
C, H, O (and N)
• Building blocks = sugars
(monosaccharide (building blocks) form polysaccharides that take on specific terms like PD)
• Ex. Polysaccharides and peptidoglycan
C: 1
H: 2
O: 1
(glucose C6H12O6)
If a sugar contains N what does that mean?
sugar derivative
- if it contains N its a sugar derivative never part of glc fructose ribose deoxyribose structures
Nucleic acids
C,H,O,N,P
• Building blocks = nucleotides
• Ex. DNA and RNA
Protein is ____% of dry weight
55
RNA is ____% of dry weight
20.5
Describe where each Nucleic acids–C,H,O,N,P are part of
C,H,O - part of sugar (ribose or deoxyribose)
N - part of nitrogenous base
P - part of phosphate group
Why does RNA make up such a high percent of the dry weight of a cell?
Bc its a precursor to proteins
transcript
Carbon is ___% of dry weight
55
backbone for all organic macromolecules
Hydrogen is ___% of dry weigh
8.2
only forms 1 cov. bond
Selenium is ___% of dry weight
<0.01
- used to form selenocysteine (an AA MODIFICATION)
- even though its small you still need it
inorganic ions
Other Macronutrients K, Mg, Ca, Fe)
- Often serve as metabolic co-factors
• Non-protein component required for enzyme function
- enzyme itself is a PROTEIN
• Enzymes involved in protein synthesis require K+
• Cytochromes (e- carriers) require Fe2+
( complexing together with iron affects reduction potential which affects e- affinity so those e- carriers will have diff. affinity (pull/desire for those e-‘s) which will correlate with amount of energy that’s released)
co- factors
like a sidekick
- can’t function without the sidekick
Describe glycolysis
glucose –> glucose 6-P
glucose –>(hexokinase - protein; has cofactor of Mg2+, allowing formation of G-6P) glucose 6-P (-)ly charged
Mg2+ (NON-protein) (+)ly charged (therefore provides temporary relief for charge repulsion that (-) charges will have with 1 another
ATP –> ADP Pi comes off and goes to form G-6P
List some other functions of inorganic ions
- Mg2+ helps stabilize membranes and nucleic acids to relieve charge repulsion
- Ca2+ helps stabilize cell walls (dipicolinic acid), and plays a role in heat stability of endospores
What needs to be considered when making a media?
**All of the macronutrients should be considered when making media
- when making a growth medium with intention tp grow something in the lab you need to be absolute certain your feeding them all these things
not all organisms like the same thing
- concentrations will be off b/c they are wrong
Are we able to grow everything in the lab?
NO
- be they are specific
Micronutrients
• Elements required in very small amounts (trace elements)
- doesn’t mean they are unimp. ( they are CRITICAL for their function)
- just b/c you don’t need a lot of something, doesn’t mean you can remove it & it still will lead to an unhindered life (absolutely critical you always have this material in the cell for full growth & success & viability)
- Usually serve as cofactors for enzymes (provide ionic stability, can’t function without them)
- Ex) Mn, Zn, Co, Ni, Cu, Mo
• Se is required to make the unusual amino acid selenocysteine(derivative of cysteine AA (has S); been modified - allowing add. function/diversity)
Describe the function of Zn 2+
Carboxypeptidase; enzyme used in digestion of protein in intestine
- uses Zn 2+ to stabilize active site of its enzyme
- anywhere you have a (-) charge with your substrate, the (+)ity can provide ionic stability or electrostatic activity
Why is C so abundant, & O and H are less?
C - can form 4 cov. bonds - excellent candidate to build lipids, carbs, AA’s & nucleotides b/c you can build elaborate castles (lots of diff. options on how they can be built)
O - can form 2 cov. bonds - less diversity/options
H - can form 1 cov. bond
Growth factors
• Small organic molecules required for growth
- CELLS NEED TO HAVE THEM TO SUSTAIN GROWTH
- things that cells can sometimes make (have recipe in the genetic material that calls for that)
• If an organism cannot synthesize the growth factor, then it must be added to medium to grow that microbe in the lab
If you have an organism that CANNOT synthesize a GF, then gets put into a growth medium that doesn’t have that GF? Will it grow?
- either they won’t grow or they’ll grow abnormally (certain pathway for ex won’t be prod.)
- depends on what it was critical for
- expect destruction of some kind
- GFs can either be made b/c organism has a recipe within its genetic makeup to actually cell for that OR it must be provided
If you have an organism who CAN synthesize the GF. Will it rather wanna make it on its own or take what you provided in the GM? You can give or not give it to him. Would he rather you give it to him or make it on its own?
- GIVE IT TO HIM –> b/c LESS energy (less time/work)
think: someone can buy you a car or you can earn it on your own
- you’ll take, but doesn’t mean you’re not capable of working for it - have opp. to do that
List the Three classes of growth factors:
- Amino acids
- Purines and pyrimidines
- Vitamins
Amino acids
• 20 amino acids are needed for protein synthesis
AA= lego block protein= lego castle
Purines and pyrimidines
- A,G,T,CandU
* Needed to make nucleotides, building blocks of DNA and RNA
Vitamins
- Small molecules used to make organic cofactors (usually something needed by an enzyme for full function)
- Non-protein components required by some enzymes(enzymes= protein)
• Ex) Nicotinic acid —> NAD+
Describe which is the cofactor and which is the vitamin
Ex) Nicotinic acid –> NAD+
Nicotinic acid - vitamin
NAD+ - cofactor, non-protein (since this cofactor is ORGANIC its called a COENZYME)
- imp. cofactor that needs to be present for glycolysis & Kreb’s cycle
- necessity to have it
- inability to produce energy if the enzyme doesn’t have it & cell will die
Compare and contrast which organisms have growth requirements vs ones that don’t be specific
• Many have no growth factor
requirements
- organism can make anything that requires by itself
- “not picky eaters”
• Ex) E. coli
- very autonomous (do a lot by themselves)
- Give basic N C and O and then leave organism alone and the will fulllfill their needs
- this is why we use E.coli so much b/c we don’t have to do much
- Addition of growth factors to medium may promote growth
- Some bacteria require many
PICKY - can’t make it on their own (CAN’T COOK)
Lipoic acid
used in intermediate step and TCA cycle
decarboxylation of pyruvate and alpha ketoglutarate
- serve as temporary electron carrier
- but if not there you screw up mechanism for these 2 enzymes and they aren’t able to produce their product which stops metabolism
- > can’t produce enough NaDH2 FADH2- can’t produce ATP
biotin
formation of c-c bonds covalent bonds
- if enzyme is forming these bonds it’ll have biotin as imp co-enzyme b/c its not easy to form covalent bonds
FUNCTION: fatty acid biosynthesis; some CO2 fixation rxns
Describe PABA (p-aminobenzoic acid) growth factor
- precursor of folic acid
- some bacteria use this pathway for folic acid syn.
folic acid
- necessary by bacterium to form AA & some nucleotides
What happens if a bacterium does not have folic acid?
- if bacterium doesn’t have folic acid, it cannot form full spectrum of nucleotides & AA’s that it req’s
What are sulfa drugs what is the function of them?
antibiotic
- analogs to PABA (look like PABA, but isn’t), which confuses bacterium & allows it in
- when you take this antibiotic bacteria get confused “it looks like PABA” but it isn’t PABA so you prevent folic acid synthesis in bacteria when this antibiotic is present in body
- if bug can’t make folic acid it can’t make amino acids and nucleotides
Why is the sulfa drug good?
- antibiotic has PERFECT selective toxicity - b/c we don’t make folic acid, we get it from our diet, therefore when we take the drug it doesn’t interfere with our own folic acid syn. b/c we don’t do our own folic acid syn
- can be resistant
Nutrient sources usually identified by ___
element
Where do they get H,O from?
- No specific nutrient
- Found in H2O and organic media components
- H20 is always part of growth medium
- organic media; ex- glc C6H1206
Where do they get P from? Why
Usually provided as phosphate salt (PO43-)
• Ex) K2HPO4, KH2PO4
• Reason: usually acquired as PO43- in the environment
* NO atmospheric source of phosphorus in the environment except in select geographic locations which have phosphine
• In freshwater systems PO43- is often limiting
- organism will run out of it so they can’t build nucleotides, lipids and ATP
Limiting nutrient
- In relatively low concentration compared to other nutrients
- When it runs out, growth stops despite other nutrients present
What becomes limiting first & second?
in our laundry detergents there is PO43- so now you have an additional source which isn’t supposed to be there so its no longer limiting meaning you won’t run out of it
nitrogen becomes limiting next
Why do ppl fertilize lawns?
limiting nutrient-> When it runs out, growth stops despite other nutrients present
- this is why ppl fertilize their lawns to add things that are missing in the soil to encourage growth
- if you add to much it’s osmotically active so it dehydrates the grass
Where do they get N from?
(many possible sources)
Inorganic N
Organic N
Atmospheric N2
Inorganic N
•no carbon attached
- Provided as salts (ex. KNO3 or NH4Cl)
- getting K and Cl requirement at the same time
• Must be reduced to NH3 – used to make amino acids (-NH2)
2NO3- –> N2 —> 2NH3
NO3-= fully oxidized (bonds to oxygen), behaves as e- acceptor N2= intermediate redox step 2NH3= fully reduced e- donor
Why can’t N2 be used by many organisms?
N-N= have a short but strong triple covalent bond
N2 makes up 78% of atm composition but b/c triple bond is so stable you need 400 degrees celsius and 200 atmp to disrupt it
-+ to use this the cell needs enzymes which provide mechanism to disrupt the bond but this uses a lot of energy so only certain organisms have capacity to use N2
Organic N
Provided as N rich organic (has C backbone) molecules
(ex. Amino acids or short peptides)
- need to break bonds to get free amino acids and you get energy when bonds are broken, breaking ordered molecules= releasing energy
- does not need to be reduced
- you want this
Provided a bug had the option to chose inorganic N and organic N which would they rather have?
the organic N b/c they don’t have to do anything
- as a researcher it is also better b/c it grows very quickly b/c it doesn’t have to pay pr work to build molecules
Atmospheric N2
- N2 is reduced to 2NH3 – Nitrogen fixation
- NH3 is used to make amino acids
• Energetically expensive
- enzymes need a lot of ATP
• Can only be done by some Bacteria and Archaea – not by eukaryotes
Rhizobium leguminosum
- capable of N fixation
- found in the root nodules of plants
- plants provide them with nutrients , photosynthetic sugars and they fix N; symbiotic relationship
- enzymes of nitrogen fixation are poisoned by O2 - photosynthesis produces O2 so they have little chambers to protect them
Where do they get S from?
Inorganic S
Organic S
Inorganic S
• Provided as salts (ex. MgSO4)
-SO4 provides you with oxidized sulfur
- Must be reduced to the level of S2- ; used to make amino acids
- Assimilative sulfate reduction
Describe SO42- –> S2- –> H2S
SO42-; fully oxidized
S2- ; intermediate redox state
H2S ; fully reduced
- the water we produce in the ETC helps meet the water demand of the cell
- more reduced form of Sulfur that is metabolic waste (can poop it out) but here it assimilates into an organic molecule ( to build things like AA)
- maintains balance in interior of the cell
What’s it called if they take reduced form of S after ETC & just poop it to outside of cell?
non assimilative sulfate reduction
What can so42-be used for?
can be used as a substitute for O2 in the ETC
- anaerobic respiration
- don’t get 32 ATP but better value then fermentation
Organic S
• Pre-made amino acids (cysteine and methionine)
- has S as framework;less energy to assimilate
• Less energy to assimilate
- can grow a little bit faster
C
Refers to the source the majority of C in macromolecules
** main component for organic molecules
DOESN”T matter if plant cell bacterial cell animal cell we all have the same core requirements for carbon
How are organisms placed into groups?
Organisms placed into 2 groups based on how they obtain C:
Heterotrophs
Autotrophs
Heterotrophs
• Use organic carbon
- carbon that’s a component of lipid, carb something that’s an organic molecule that contains carbon in its framework,
• One or more C is reduced (ie. a C atom with one or more H’s)
- lots of energy can come from stripping these e- from the framework and using them in redox reactions to generate ATP
• Ex) Organic acids (lactic acid), alcohols (OH), carbohydrates, amino acids
- energy rich if they get catabolized
Autotrophs
• Use inorganic carbon (CO2) as their sole source of carbon
- “can cook”
- take carbon source and assimilate it into a larger structure
- CO2 comes from respiratory system all animals that do respiration this way, burning fossil fuels, fireplaces
- Requires energy to assimilate
- Photosynthesis
” molecules are gaseous energetically happy and disordered the way they are and then they get fixed into this structure and energy is stored so that when you break it later you can obtain energy
• Ex) Anabaena
Defined medium
• Exact chemical composition is known
- we know exactly what is in the medium b/c we follow a recipe and add ingredients one by one
• Useful for studying metabolism
- can change up a factor
ex; add extra stuff take away stuff and see how it affects growth
- reproducible- publish in an article
Minimal medium
• A defined medium that provides the minimum nutritional requirements for growth (ie. No growth factors)
- not super rich in nutrients just provides the bare minimum in order to promote survival
Complex medium
• Exact chemical composition is not known
- “ going to the store and buying pre- made pizza” you have no idea what’s exactly inside
• Often made from meat or yeast extracts
- meat= animal tissue made of lipids nucleic acids carbs
- yeast extracts= mashup of yeast to make an extract
- can also add blood which is a growth medium and what we use to sustain our cells and grow
- but you will never be able to give exact composition of blood we know it has oxygen iron and protein
- Supply a variety of growth factors
- Ex) T-soy broth and plates
What does t- soy broth not have?
Agar so plate should have agar
Differential medium
- Allows different bacteria to be distinguished
- Ex) Blood agar – T-soy plate (nutrient medium) + 5% sheep’s blood (allows you to test for hemolysis)
- Allows differentiation of hemolytic bacteria
What is hemolysis?
blood cells that split
a-hemolysis
incomplete destruction of blood cells
b- hemolysis
completedestruction
g-hemolysis
–no destruction
- organism is growing but can’t break blood cells apart
MacConkey
ex of a differential medium
includes a ph indicator in growth medium
- any acid reacts= pink
- non acid waste= white
type of metabolism determines what it will look like
Selective medium
Contain ingredients that inhibit the growth of unwanted microbes
• Allow only specific microbes to grow
( # of microbes that can grow decrease, becomes more manageable, add something thats toxic)
- if you took a sample of soil and put it on a petri dish you wouldn’t be able to analyze a specific microbe b/c there is to much growth , to much present
Mannitol salt agar
- ex of a selective medium
• Contains very high salt, so that only halotolerant
bacteria will grow
- most organisms under hypertonic conditions that salt creates become dehydrated and lose ability to maintain H20 and survive only the ones that can manage water conditions will grow
• Used to isolate staphylococci from skin
- b/c all other organisms that can’t tolerate this won’t grow
- ex- S. aureus
Bile salts
can add bile salts to make a medium selective
- > bile salts are used in intestine to emulsify or rip apart fat to make a hydrophobic molecule easier to digest
- > organisms are made out of lipids so it’s very difficult for them to handle conditions in the intestine, so only organisms who are built to tolerate intestine b/c they grew up their will do so everything else will be selected against
If you want to grow intestinal bugs what should you do?
throw in some bile salts to get ride of organisms that aren’t b/c most will be destroyed as a result of this
Enriched medium:
- Supplemented with special nutrients to encourage the growth of fastidious bacteria
- Complex nutrient requirements – require many growth factors
- Ex) Blood agar (encourages growth), chocolate agar (nutrient source)
Can ALL organisms be grown in the lab?
NO most cant
- we use metagenomic analysis to identify them
( non- culturable)
Metabolism
The sum total of all of the chemical reactions that occur in a cell
Catabolic reactions (catabolism)
Energy-releasing metabolic reactions (e.g. fermentation, respiration)
- break down molecules
- glc gets broken down in respiration- results in 6 CO2
- like a paycheck
- creates disorder= energy is released b/c everything wants disorder “things are happier”
Anabolic reactions (anabolism)
Energy-requiring metabolic reactions (biosynthesis)
- things you need to pay for
- create order; things don’t like to be ordered
- building RNA proteins
Every single LIVING organism can be categorized by 2 things:
- carbon source
2. energy source
Microorganisms grouped into energy classes depending on their source of electrons and energy:
- Chemorganotrophs
• Energy from chemical reactions involving
organic material - Chemolithotrophs
• Energy from inorganic chemical reactions - Phototrophs
• Energy from light
Chemorganotrophs
Energy from chemical reactions involving organic material - this is us - chemotrophs - C6H12O6=lots of e-, lost of energy available if you break them down
Chemolithotrophs
Energy from inorganic chemical reactions
- H2 S
How much energy is avail. in H2S as compared to C6H12O6?
less
What will the compensation be b/t an organism that uses glucose to supply itself b/c its a chemorganotroph & an organism that is chemolithotrophic & uses H2S to supply its energy?
Eat more H2S to get same amount of energy that glc provides
-> process more to get same energy yield
Phototrophs
Energy from light
Microorganisms grouped with respect to carbon source:
- Heterotrophs
• Use organic carbon for building cell carbon
and biomass - Autotrophs
• Use CO2 to synthesize cell carbon
• a.k.a. primary producers
What about if an organism uses palmatic acid (C16 fatty acid) for C & energy. What is its classification?
chemoorganoheterotroph
What about if an organism uses light energy & organic chemicals. What is its classification?
photoheterotroph
What are we in term of our chemical & nutrient requirements?
chemoorganoheterotroph
-> most medically relevant bacteria are this
