Microbiology Exam 2
Last Modified: 2012-02-23
ATP is made up of
- a ribose (which is a sugar)
- three phosphate group
the collision energy required for a chemical reaction which is the amount of energy needed to disrupt the stable electronic configuration of any specific molecule so that the electrons can be rearranged.
Energy of Activation- given for the energy required to convert reactants into products in a chemical reaction
sequences of chemical reactions
They are determined by its "enzymes" which are in turn determined by the cell's genetic makeup
- they are proteins (globular), produced by living cells, that catalyze chemical reactions by lowering the activiation energy without increasing the temperature or pressure within a cell.
- They can operate at a low temperature. Each has a unique 3D shape.
- They end in "- ase". Enzymes are characterized specifically which is a function of their active site. There are SIX classes of enzymes.
- The transition of reactants to products requires a certain amount of energy.
- Derived from Vitamins
- Enzymes increase the rate of chemical reactions.
- Make it easier for cells to produce the products of various chemical reactions in the cell
- By accepting atoms removed from the substrate or by donating atoms required by the substrate.
- Also some enzymes act as electron carriers, removing electrons from the substrate and donating them to other molecules in subsequent reactions.
- Nicotinamide adenine dinucleotide (NAD+) involve primarily in catabolic (energy-yielding) reactions
- Nicotinamide adenine dinucleotide phosphate (NADP+) involved in anabolic (energy-requiring) reactions.
both function as electron carriers.
- flavin adenine dinucleotide (FAD) an electron carrier
- coenzyme A (CoA) - this enzyme plays an important role in the synthesis and breakdown of fats and in a series of oxidizing reactions called the Krebs Cycle
- Substrate concentration- increase substrate concentration increases rate. High substrate concentration = enzyme saturation, that is the active site is always occupied by substrate or product molecules.
- pH- must maintain optimal pH, if not causes denaturation.
- Temperature- must obtain optimal temp.. too high or too low causes the protein to undergo denaturation (breakage of the noncavalent bonds such as hydrogen bonds)
- Inhibitors- way to control growth of bacteria by controlling the enzymes
Competitive- fill the active site of an enzyme and compete w/ the normal substrate for the active site. They do not form a product. Some bind permanently. Some bind & leave, slowing the enzyme's activity.
Noncompetitive- also called allosteric ("other space"). The inhibitor binds to another site on the enzyme other than the substrate's bindng site, called the allosteric site (prevents cell from making excess substances). This causes the active site to change shape making it nonfunctional.
- Oxidation-Reduction Reaction (often called "Redox" Reaction)
- ATP Generation
Much of the energy released during oxidation-reduction reactions is trapped within the cell by the formation of ATP. Specifically a phosphate group "P" is added to ADP w/ the input of energy to form ATP. The addition of "P" to a chemical compound is called Phosphorylation.
(When this "P" is removed, energy is released)
- Substrate-level phosphorylation- ATP is usually generated when a "high energy P is directly transferred from a phosporylated compound PO4 -(substrate) to ADP.
- Oxidative Phosphorylation- transfers electrons from org compounds to electron carriers (NAD+ & FAD). They use the Electron Transport Chain (ETC)
- Photophosphorylation- occurs only in photosynthetic cells which contain light-trapping pigments as clorophylls (release electrons which are oxidized & used to generate ATP.
Eukaryotes in the inner mitochondrial membrane
Prokaryotes in the plasma membrane
1 step: Glycolysis (also known as Embden-Meyerhof pathway)
2nd Step: Krebs Cycle (also called TCA- tricarboxylic acid cycle or citric acid cycle)
3rd Step: Electron transport Chain
the glucose rearranges & now another ATP is invested & another phosphate is added to glucose completely changing the structure. By adding extra groups the compound becomes unstable. By investing the two ATP's, the two phospate groups at the end of the compound has broken it apart into two pyruvic groups called G3P molecules.
Pentose phosphate pathway
Entner- Doudoroff pathway
- uses pentose and NADPH
- operates w/ glycolysis
- produces NADPH and 1 ATP
little less energy
- Produces 2NADPH and 1 ATP
- DOES NOT involve Glycolysis
- typically find in gram negative bacteria such as Pseudomonas, Rhizobium, Agrobacterium
occurs instead of glycolysis
To the ETS to drop off their electrons!!
NOW ITS TIME FOR THE BIG PAYOFF (LOTS OF ATP TO BE MADE)
- Occurs in mitochondrion inner membrane of Eukaryotes & plasma membrane for prokaryotes
- Series of redox reactions involving electron transporters (cytochromes)-- in aerobes (type of respiration uses oxygen), final electron accepter is oxygen (-) which combines w/ 2 protons, hyrdrogen H+) to make water H2O.
- Energy released from these reactions is used to set up a proton (H+) gradient across the membrane (pumps protons = active transport)
- ATP is made by oxidative phosphorylation (chemiosmosis)
Pathway: ATP produced NADH produced FADH2 produced
Glycolysis 2 2 0
Interm step 0 2 0
Krebs Cyc 2 6 2
TOTAL 4 10 2
Each NADH produces ** 3 ATP **
Each FADH2 produces ** 2 ATP **
By substrate level By Oxidative Phosphorylation
Pathway: phosphorylation FROM NADH FROM FADH2
Glycolysis 2 6 0
Interm step 0 6 0
Krebs Cyc 2 18 4
TOTAL 4 30 4 TOTAL = 38 ATP
** 36 ATPs are prod in Eukaryotes
- Occurs in the absence of oxygen.
- the final electron is an inorganic compound rather than oxygen O2.
- The inorganic compounds example are NO3, SO4. CO3
- Usually a shortened Krebs Cycle & ETS, so not as many ATP are produced.
- Yields between 2 and 38 ATPS.
Electron acceptor Products
NO3- NO2-, N2+, H2O
SO4- H2S+, H2O
CO32- CH4+, H2O
- Releases energy from oxidation of organic molecules
- Does not require oxygen
- Does not use Krebs Cycle or ETC
- Glycolysis is only ATP producing step
- Uses an organic molecule as the final electron acceptor
- Yields 2 ATP
Produces ethyl alcohol + CO2
- Produces Lactic Acid
Two important genera of lactic acid bacteria are Streptococcus and Lactobacillus are microbes produces ONLY lactic acid... there are called Homolactic
Organisms that produce lactic acid as well as other acids or alcohols are known as heterolactic
Microbes can oxidize substances other than glucose such as lipids and proteins through two processes:
- Lipid Catabolism
- Protein Catabolism
Lipid Catabolism- microbes uses an enzyme called "lipase" to break down fats (hydrolyzed) into glycerol and fatty acids.
- Glycerol (3-carbon) is converted into PGAL (G3P) also a 3-carbon
- PGAL then enters respiratory pathway
- Fatty acids are broken down (hydrolyzed) into 2-carbon fragments by "beta oxidation"
- Each 2C fragment is converted to Acetyl
(multi C fatty acid -------> 2C fragment -------> 2C Acetyl)
- Acetyl then enters the respiratory pathway
Proteins are TOO large to pass unaided thru the plasma membrane. Microbes produced enzymes- protease & peptidase to break down proteins into amino acids to pass thru. The amino acids are "deaminated"- the removal of an amino acid group from an amino acid to form ammonia NH4+ which will be excreted from the cell, the remaning organic acid (keto acids) can center the Krebs cycle.
The keto acids are Pyruvic acid, Acetyl and Intermediates
Proteins, Carbohydrates and Lipids can all be sources of electrons & protons for respiration. These food molecules enter Glycolysis or Krebs cycle at various points.
Proteins break down to amino acids can enter at glycolysis, Acetyl CoA or Krebs
Carbohyrdates breaks down to sugars and enter at Glycolysis
Lipids break down to two parts either glycerol that enters at Glycolysis or Fatty acids that enter at Acetyl CoA.
- phototrophs - uses light as their primary energy source
- chemotrophs- depend on oxidation- reduction reactions of inorganic or organic compounds for energy
- Photoautrotrophs- light as energy source, use CO2 as carbon source. Ex: photosynthetic bacteria, algae & green plants
- Photoheterotrophs- light as primary energy source, organic compounds as carbon. Ex: Chloroflexus, Rhodopseudomonas.
- Chemoautotrophs- inorganic compounds as source of energy. Use CO2 for carbon. Ex: Nitrosomonas, Hydrogenomonas
- Energy & Carbon source the same!
- Grouped by source of organic carbon
- Saprophytes- live on dead matter,
- parasites- derive nutrients from living host
** MOST MEDICALLY IMPORTANT MICROBE GROUP**
- it is the study of heredity
- transmission of biological traits from parent to offspring
- expression & variation of those traits
- function of our genetic material (DNA) is to make proteins
- structure of our genetic material
- how this material changes b/c it does change through the process of transcription and translation
Organism level- the genotype & phenotype
Chromosome level (Eukaryote only)
the processes of replication, transcription and translation
DNA replicates, they transcribe and translate and that is how DNA goes from being a blueprint to functional.
All the genetic information in a cell.. your genes, your dna, your genetic makeup
for prokaryotes- plasmids make up part of the genome for prokaryotes. Plasmids allow bacteria cell to be anabolic resistant.
structure containing DNA that physically carries hereditary information; the chromosomes contain the ALL OF YOUR GENES!
The genes are found
a segment of DNA that encodes a functional product, usually a protein (viruses can have RNA)
There are 3 types of gene:
- structural genes- these genes make protein
- Genes that Code for RNA
- Regulatory genes
the "expression" of those genes.
- They can change depending on the expression of the genotype
Prokaryotes by enzymes coiled tight bundle by enzyme called gyrase is a topoisomerase
For Eukaryotes we have to fit inside the nucleus so it will requires alittle work to fit 6ft info inside the nucleus so we will also use enzymes and a specific protein:
- wound around a "histone proteins"... 4 of these form nucleosomes
- Nucleosomes condense, coil into chromatin fibers
- Chromatin fibers "supercoils" and condenses into chromosomes
- Nucleic acids are the polymers & Nucleotides are monomers
- Nucleotides going to have a 5-carbon sugar. If DNA it's deoxyribose and if RNA its Ribose. Both have a phosphate group & a nitrogenous base. The bases differ.
- Purines: A+G (adenine & guanine) 2 H+ bonds
- Pyrimidines: C, T & U (cytosine, thymine, uracil) 3 H+ bonds
- The bases pair by A----T and G----C (think of George Clooney)..
- RNA "DIFFERS" RNA replaces T w/ U, so the pairing for RNA is A--- U, G--- C (G----C never changes)
- 2 strands twisted into a double helix
- On each side of strand, we have a sugar, a phosphate group, nitrogenous base. For it to line up just right .
- nitrogenous bases are where hydrogen bonds form! (think of the latter! the steps are the nitrogenous bases that connects the sides (the sugars & the phosphates)
- Each strand is parallel (5' to 3' & 3' to 5') & provides a "template" for the exact copying of new strand
- Order of bases constitues the DNA code which gives our language (A, G, C, T, U)
- Maintenance of code during reproduction. We have to make sure we get the same thing. Constancy of base pairing guarantees that the code will be retained! A-----T and G-----C.
- Providing variety. Order of bases responsible for unique qualities of each organism (gene sequence). the possible arrangement of nucleotides is nearly infinite: 4n = #Nucleotides. So for a 1000 bp genes the combinations are 41000
that is alot!!
- Gregor Mendall- the "Father of Genetics" monk & experimented w/ Pea plants.
- Avery, MacLeod & McCarty- showed DNA was the molecule carrying the blueprint for life. Nobel prize.
- Erwin Chargaff- components of DNA- the base pairs A, G, C, T, U. You always have the same amount of each. 64 adenines then you have 64 Thymines
- Maurice Wilkins & Rosalind Franklin(woman)- XRay crystallography gave clue to double helix structure
- Watson & Crick- structure of DNA b/c of Rosalind xray. Nobel prize
- Due base pairing, each strand serves as a template for the synthesis of a new strand.
- DNA replication is semiconservative b/c each chromosome ends up w/ one new strand of DNA & one old strand. Only 1/2 of the DNA is going to be new the other 1/2 is the template.
Once aligned, the newly added nucleotide is joined to the growing DNA strand by an enzyme called DNA polymerase (proofreader) .
- The parental DNA is unwound a bit further to all the addition of the next nucleotides. The point at which replication occurs is called the replication fork. As the replication fork moves along the parental DNA, each of the unwound single strands combines w/ new nucleotides.
- The original & new strand rewind back & is put back together by the enzyme Ligase.
Looking at the structure of the DNA, the paired DNA strands are oriented in opposite directions relative to each other. The 5' end is always the end of the phosphate group & the 3' end is always the end with the hydroxyl group. DNA polymerase can only add nucleotides from the 5' to 3' end. Leading strand (5' to 3') is synthesizing continuosuly but the lagging (3' to 5') strand is synthesized discontinuously. For DNA polymerase to work here, it has to "jump forward" so it can work backwards.
When this happens it creates "gaps" or "fragments" called Okazaki fragments. Because your DNA polymerase is always jumping ahead you have sections not put together.
We cannot leave it like this b/c the copy strand has to be like the original strand. So the DNA ligase is brought in to patch the holes. It will get "rid" of the Okazaki fragments. It only needs to do this on the lagging strand (3' to 5')
This is how it works in the prokaryotes (cytoplasm) & Eukaryotes (nucleus) cells
DNA (info) ---------------> RNA -----------------------> PROTEINS
(RNA Polymerase) (Ribosomes)
Reference books in the library are full of info but you cant check them out. You can copy what you need out of the reference book. DNA is like a giant cookbook, its full of recipes but we want a specific one on chocolate cake. We find that recipe, make a copy, bring home and make out chocolate cake
- messenger RNA (mRNA)- is the message, it can leave the nucleus. It is the copy of the DNA
- transfer RNA (tRNA)- this will go and get what we need.
- ribosomal RNA (rRNA)- found in ribosomes w/ subunits: rRNA and a protein. rRNA acts like a reader. It will tell us what amino acids we need.
- Single stranded (except in some viruses)
- Uses Uracil (U) instead of Thymine (T)
- Ribose is the sugar
- still going to have the same 5' to 3' strand. but using fructose instead of glucose
- RNA is synthesized from DNA template
- Nucleus of Eukaryotic cells
- Cytoplasm of prokaryotic cells
- RNA polymerase binds to DNA at a promotor site (starter) and begins "scanning" the gene.
- RNA polymerase will bring in free RNA Nucleotides in & attached to their complimentary nucleotides on the DNA strand by Ribose (sugar)
- RNA synthesis also progressess 5' to 3'
- mRNA synthesis ends when the RNA polymerase reaches the last triplet of nucleotides called the terminator. When its done copying it curls up
- Newly transcribed mRNA is then released from the DNA strand. What you created is the mRNA
- We have junk DNA like fillers (introns)! We want the real info (exons)
- Exons- segments of gene that code for proteins
- Introns- segments of a gene that do not code for proteins.
- RNA is processed by ribozyme - removes the introns from mRNA and splices the exons together
Eukaryotic DNA contains segments that do not code for proteins
- mRNA copy of a DNA gene is "read" by a ribosome. Ribosomes are the site of protein synthesis. What we are trying to make is the protein.
- The mRNA is a copy of a DNA gene (instructions for making a protein)
- Occurs in cytoplasm of both prokaryotes & eukaryotes
** transcripton & translation can occur at the same time in prokaryotes b/c the processes are not separated by a nuclear membrane.
*** but not in Eukaryotic cell b/c transcription has to take place 1st in the nucleus then translation
- Ribosomes "read" the genetic message 3 nucleotides at a time.
- The triplets of nucleotides are called codons - responsible for a set code of amino acid
- Each codon codes for one amino acid
- The trick with codons is you have 64 codons.
- 61 of them code for amino acids. But there are only 20 amino acids. This implies that "some" of our codons will code for the same amino acid.
- The remainder of the 3 are the 3 nonsense codons (stop codons) Where to start and where to stop
- The genetic code is degenerate. This helps in case if we get a mutation in our dna, the amino acid may be code
- Translation of mRNA begins at the start codon. Its always AUG (easy to remember.. the month we start school is August)
- Ribosome slides along the mRNA, reading each codon (tells the sequence) it tells what we need
- Amino Acids are brought to the mRNA molecule by tRNAs
- Each tRNA has an anticodon to complement
- Translation ends when the ribosome reaches the end the 3 terminator codons (UAA, UAG, UGA)
** Some transcribed genes arent translated. tRNA & rRNA, RNA primers serve add't functions**
Most genes are we refer to as Constitutive genes.
- Constitutive genes are NOT regulated- always "on" always working (60% of our genes are). These genes are active & continously making enzymes or other proteins that are needed constantly
- Products are produced at a fix rate.
- Regulatory genes you can turn on/off...
- Induction ("turns on")- the default is to be off
- The presence of a substance in a cell induces transcription of enzymes related to that substance.
- Ex: Iac operon
- Repression ("inhibits") turns off- so the default is to be turned on
- An excess of end products not being used stops transcription of enzymes related to that substance
- Ex: tryp operon
a coordinated set of genes, all of which work together. Found in prokaryotes
2 types- based on regulation
- Inducible operon- operon is turned ON by substrate (default to be turned off) This will be used to help regulate transcription.
- Repressible operon- operon is turned OFF by the product synthesized (default to be turned on)
- any permanent, inheritable change in DNA
- alterations of the nucleotide sequence (ATGC) bases
- involves either loss, addition or rearrangement of base pairs (can be multiple)
- Spontaneous mutation- random, due to during replication error
- Induced mutation- from exposure to mutagens (physical, chemical; disrupts DNA). Induced mutations are what we can control!
- Mutations may be:
- neutral (silent- meaning the outcome will not be affected)
- Mutation caused by the addition or deletion of one more bases
- Shifts the "translational reading frame" (remember codons are always 3 letters.. so if we add in an extra bases pair.. we have to shift everything over)
Ex: THE CAT ATE THE RAT
Frameshift: TTH ECA TAT ETH ERA T..
Nonsense Mutation- a base substitution resulting in a nonsense codon.
- By creating a nonsense (stop) codon in the middle of an mRNA molecule, some base substitutions effectively prevent the synthesis of a complete functional protein
Missense mutation- this mutation will cause a change in the DNA, if the base substitution results in an amino acid substitution in the synthesized protein.
example: sickle cell anemia
- Mutations dont occur as often as we think
- Spontaneous mutation rate = 1 in 106 replication of DNA
- Induced Mutations rate = 1 in 104 replications of DNA
- Rates can be "altered" by the presence of mutagens
- Any agent in the environment that brings about a mutation
- Includes chemicals & radiation
- Look at the structure of the normal nitrogenous base of our nucleoside Adenine. (The golden rule of DNA, you dont have to worry b/c (A--->T) and (G---->C)
- Beside this is a nucleoside analog. These analogs mess up how DNA is suppose to base pair. They sorta of trick DNA, in that its a different base. So instead of Adenine it creates Aminopurine.
This examle of a chemical mutagen
Can be identified by selecting or testing for an altered phenotype. Two Detection Methods are used.
- Positive (direct) selection- detects mutant cells b/c they grow or appear different. Look for the growth or presence for organism that are "resistant" to antibiotics. If its resistant to antibiotics, its had a mutation in dna to grow in the presence of antibiotics.
- Negative (indirect) selection- detects mutant cells b/c they do not grow. Use a technique called "replica plating".
To identify the capability of synthesizing amino acid "histidine".
- 1st - you have master plate w/ medium with histidine.
- Using the sterile velvet press, press on the master plate that contains the grown colonies.
- You press on the plate containing histidine & you press on the plate that lacks the histidine. Incubate both.
- Growth on plates are compared. A colony that grows on the med w/ histidine but doesnt on the lacking one is auxotrophic (histidine-requiring mutant)
mutants that cause cancer in animals, including humans.
No all mutations result in carcinogens but alot of them do.
Name after Bruce Ames who invented this test.
This test uses bacteria as carcinogens indicators. The bacteria that is used often as the standard is Salmonella.
- Exchange of genes between two DNA molecules this creates recombinant dna
- Crossing over- occurs when two chromosomes break and rejoin.. this allows recombinant dna
This occurs through:
- Vertical gene transfer: occurs during reproduction between cells, passed from organism to offspring. Ex: from parent to offspring
- Horizontal gene transfer- transfer of genes between cells of the same generation. Humans can experience this through via viruses!
Ex. sister to sister
- All cells cannot undergo through transformation. Cells that "can" go through transformation are called "competent"
- Genes are transfer from bacterium to another as "naked (dna fragments)" DNA
- Occurs naturally in Bacillus, Haemophilus, Neisseria, Acinetobacter, Streptococcus & Staphylococcus... these are called competent
- Frederick Griffith -the 1st demos transformation w/ 2 strains of Streptococcus pneumoniae- one virulent (pathogenic) has a capsule, the other avirulent- lacks capsule.
- transferred from one bacterium to another. This is mediated by a plasmid (assessory to dna)
- Differs from transformation b/c it req. direct cell-to-cell contact. Must be of opposite mating type.
- Has a fertility plasmid (F factor) can be F+ or F-.
- if you are (F+) gene that allows you to produce conjugative pilus
- if "F-" then you DO NOT produce the conjugative pilus.
- F+ plasmid will replicate itself, have 2 copies. Finds "F-" and gives it a copy. The "F-" will now become (F+)
- By a "helper" in this case its by bacteriophage (virus). Bacteriophage is a virus that "attacks" bacteria. In transduction the virus is doing the work for us.
- Can occur in two different ways:
- Generalized transduction- start off w/ the prokaryotic dna. Then along comes your bacteriophage it attaches & injects its dna in the cell. The dna incorporates w/ the cell's genome. Viruses are like pirates, hijack the cell & make it do what it wants.
- Specialized transduction (not show in notes)
- Conjugative plasmid- carries genes for sex pili and transfer of the plasmid
- Dissimilation plasmids- Encode enzymes for catabolism of unusual compounds.
- We can use these plasmids & munipulate to do what we want
R factors- encode antibiotic resistance.
- Barbara McClintock discovered this in corn
- are small segments of DNA that can move from one region of DNA to another. Basically move anywhere they want!! Even move to other organisms
- This can be exciting but also scary b/c they can move in viruses!!
- Contain insertion sequences for cutting and resealing DNA (transposase)
- Complex transposons carry other genes
- Mutations and recombination provide diversity
- Fittest organisms for an environment are selected by natural selection
** The word fit means your lifetime reproductive output! **
- Two goals in life it to survive and reproduce
Words From Our Students
"StudyBlue is great for studying. I love the study guides, flashcards, and quizzes. So extremely helpful for all of my classes!"
Alice, Arizona State University
"I'm a student using StudyBlue, and I can 100% say that it helps me so much. Study materials for almost every subject in school are available in StudyBlue. It is so helpful for my education!"
Tim, University of Florida
"StudyBlue provides way more features than other studying apps, and thus allows me to learn very quickly! I actually feel much more comfortable taking my exams after I study with this app. It's amazing!"
Jennifer, Rutgers University
"I love flashcards but carrying around physical flashcards is cumbersome and simply outdated. StudyBlue is exactly what I was looking for!"