Exam 2 Flashcards
1
Q
Extremophiles
A
Microbes that can grow in extreme environments
2
Q
Solutes that decrease the availability of water to microbes (decrease water activity (aw))
A
Salts, sugars
3
Q
Hypotonic Environment
A
Low extracellular solute concentration (water wants to move in)
Ex. Freshwater lakes, streams
4
Q
Isotonic Environment
A
Same solute concentration in and out of cell, used to observe protoplasts (cell walls degraded with lysozyme)
5
Q
Hypertonic Environment
A
High extracellular solute concentration (water wants to move out), low (aw)
Ex. Dead Sea, Great Salt Lake, Peanut Butter
6
Q
Halophiles
A
Require high salt concentrations to grow
7
Q
Osmotolerant
A
Grow over a wide range of (aw)
Ex. Staphylococcus (salt-tolerant commensal of human skin)
8
Q
Mannitol Salt Agar
A
Medium used to select for Staphylococcus growth
* Pathogenic S. aureus ferments the agar
* S. epidermidis does not
9
Q
Xerophile
A
Prefer low (aw), dry conditions
Ex. Cronobacter (infant formula shortage cause)
10
Q
Compatible Solutes
A
How microbes survive in highly concentrated environments
Ex. KCl, choline, some amino acids
11
Q
Nutrients
A
Substances used in biosynthesis and energy release, required for growth
12
Q
95% of microbial cell dry weight is made up of a few ingredients:
A
Macronutrients and micronutrients
13
Q
Macronutrients
A
a.k.a. Macroelements, required in large amounts
Ex. C, O, H, P, N, S, Fe
14
Q
Micronutrients
A
a.k.a. Trace Elements, required in small amounts
Ex. Cobalt, copper, zinc, manganese
15
Q
Nitrogen Fixation
A
Reduce N2 to ammonia (NH3)
Carried out by:
* Rhizobium - commensal with plants
* Azotobacter - free living in the soil
16
Q
Microbial nitrogen sources
A
Microbes can use ammonia (NH3) or nitrate (NO3), a few can use atmospheric nitrogen gas (N2)
17
Q
In order to sustain exponential growth, food must enter cell:
A
- At high rates
- Across membranes
- In a selective fashion (non-toxic)
- Often against concentration gradient
18
Q
Passive Transport
A
Moves material from high to low concentration
* No energy
* Passive Diffusion: Only small molecules and certain gases
* Facilitated Diffusion: Uses membrane carrier proteins (provides selectivity), ex. aquaporins
19
Q
Carrier Saturation Effect
A
Only a finite number of transport proteins, once all are saturated the rate of transport plateaus
20
Q
Active Transport
A
Moves nutrients against the gradient
* Requires energy (from ATP or PMF)
* Either Primary or Secondary
21
Q
ABC Transporters
A
ATP-Binding Cassette (ABC)
* Found in all domains of life
* Solute-binding protein engages with nutrient in the periplasmic space
* Conformational change in transporter leads to nutrient transferred across the channel (requires ATP)
22
Q
Uptake ABC
A
Moves nutrients into the cell
23
Q
Export ABC (“Multi-drug efflux pumps”)
A
Move substances out of the cell
* In bacteria, mechanism of antibiotic resistance
* In animal cells, mechanism of chemotherapy resistance
24
Q
Secondary Active Transport
A
Uses potential energy of ion gradients
Ex. Lac Permease membrane protein
* Uniporter, Antiporter, Symporter mechanisms
25
Group Translocation
Method of active transport, chemically alters the nutrient
* Energy from phosphoenolpyruvate (key intermediate in glycolysis) attaches P to sugars
Ex. Phosphotransferase system in bacteria
26
Iron Uptake Problem
All microbes require iron (Fe), however there is little available outside of insoluble ferric form (Fe3+)
27
Iron Uptake Solution
Microbes release **siderophores** to acqurie Fe
* Low molecular-weight compounds
* Unique to each microbe
* Siderophore-Fe complex then transported into cell using ABC transport system
Ex. Enterobactin in E. coli
28
Metabolism
All chemical reactions within a cell, comprised of catabolism and anabolism
* Requires **enzymes** and **ATP**
29
Catabolism
Breakdown of complex molecules into smaller ones with the release of energy
30
Anabolism
Utilize products of catabolism (ATP and reducing power (NADPH)) to form new cell components and structures
31
Adenosine Triphosphate (ATP)
The main energy currency of cells due to the high free energy change of removing a phosphate, composed of:
* Adenosine: Comprised of nitrogenous base Adenine and 5C Ribose sugar
* 3 Phosphate groups
32
Aerobic/Anaerobic respiration both produce ATP by ___
oxidative phosphorylation
33
Fermentation
ATP by substrate level phosphorylation **only**
34
Photosynthesis
ATP by photophosphorylation
35
Energy-Generating Systems
* Aerobic respiration
* Anaerobic respiration
* Fermentation
* Photosynthesis
36
Redox Reactions
Used in metabolic processes to form ATP
* Electrons move from donor to acceptor
* Often involves transfer of H+ proton (ex. NAD+ becomes NADH)
* **OIL-RIG!**
37
Malate Dehydrogenase
An enzyme found in the Citric Acid cycle, utilized to oxidize malate to form oxaloacetate intermediate and reduce NAD+ to NADH
38
*Rhodoferax metabolins*
Psychrophilic, obligate anaerobe, oxidizes acetate, reduces iron
39
Ribozymes
Catalytic RNA molecules
40
Metabolic enzymes act on ___, convert to products, lower ___
substrates, activation energy
41
Reduction Potential (E0)
Equilibrium constant for redox reactions, represented by a ladder
* More negative E0 = better donor
* More positive E0 = better acceptor
* Greater difference in coupled pair releases more energy
42
Electron carriers for redox reactions divided into two groups (locational):
1. Freely Diffusable in cytoplasm (ex. NAD+ and NADPNAD+), reduced forms (NADH and NADPH) are "reducing power" of the cell
2. Membrane-bound (ex. flavoproteins, cytochromes, quinones)
43
Microbes transfer energy by moving electrons from:
Reduced food molecules (glucose), diffusable carriers in cytoplasm, membrane-bound carriers, O2, metals, or oxidized forms of N and S
44
Autotrophs
"Primary producers"
* CO2 as C source (plants, many microbes)
* Synthesize organic compounds used by heterotrophs
45
Heterotrophs
* Reduce preformed organic compounds as C source (animals, many microbes)
* Convert large amounts of C to CO2
46
Energy Sources
Phototrophs: Sunlight
Chemotrophs: Oxidize inorganic chemical compounds for energy
47
Electron Sources
Lithotrophs: Inorganic molecules as electron donors
Organotrophs: Use organic molecules as electron donors
48
Aerobic Respiration
Catabolic process that can break down an organic energy source to CO2:
1. Glycolysis
2. Citric Acid Cycle
3. ETC with Oxygen as final electron acceptor (produces a lot of ATP)
49
Breakdown of glucose to pyruvate in bacteria
3 Pathways:
1. Embden-Meyerhof (Glycolysis)
2. Pentose Phosphate
3. Entner-Doudoroff
All occur in the bacterial cytoplasm
50
Glycolysis
* 6C Stage: Glucose phosphorylated twice (requires ATP), generating fructose 1,6-bisphosphate
* 3C Stage: Fructose 1,6-bisphosphate split into 2 glyceraldehyde 3-P then converted to pyruvate
Oxidizes NADH, substrate-level phosphorylation (**net yield** 2 ATP, 2 NADH, 2 pyruvate)
51
Glycolysis NADH and ATP generating steps
Reaction 1: Glyceraldehyde-3-phosphate oxidized and phosphorylated (generates high-energy P bond, reduces NAD+ to NADH), forming 1,3-bisphosphoglycerate (**G3P dehydrogenase**), **3GP kinase**
Reaction 2: Phosphorylation of ADP by high energy metabolic substrate, generates ATP by substrate-level phosphorylation (catalyzed by **pyruvate kinase**)
52
Pentose Phosphate Pathway
* Starts by converting Glucose-6-P to Ribulose-5-P (pentose)
* Generates many sugars for biosynthesis
* **Yields 6 NADPH** (reducing power for biosynthesis) and 1 ATP
53
Entner-Doudoroff Pathways
* Combines reactions of glycolysis and pentose phosphate
* Net 1 ATP, 1 NADH, and 1 NADPH
* Important for growth of *Escherichia coli* in intestine
54
Citric Acid (TCA) Cycle
* Pyruvate completely oxidized to CO2
* In cytoplasm of bacteria
* Generates: CO2, numerous NADH and FADH2, precursors for biosynthesis, GTP
55
Electron Transport Chains
* Electrons from NADH and FADH2 generated by glycolysis and TCA cycle are transferred through a series of membrane bound electron carriers to a terminal electron acceptor
* Electrons flow from carriers with more negative E0 to more positive, energy is released to make ATP by oxidative phosphorylation
* 3 ATP can be generated per NADH using O2 as terminal acceptor
56
Chemiosmotic Hypothesis
Oxidative phosphorylation hypothesis posed by Peter Mitchell
* Energy released during ETC establishes proton gradient and charge difference across the membrane, source of PMF
57
PMF drives ATP synthesis:
Electron flow in ETC causes protons to move outward across membrane, ATP made when they move back in via **F1F0 ATP synthase**
58
F0 Subunit
Proton channel, ring of C subunits that rotates
59
F1 Subunit
Gamma shaft rotates, conformational changes in sphere of alpha and beta subunits within cell leading to ATP synthesis
60
*Shewanella*
Gram - bacterium, can transfer electrons *extracellularly* onto metals ("breathes metal")
61
A microbial fuel cell ___
Can capture extracellular electrons to generate electricity
* Low O2 conditions (anoxic) promotes formation of electrode biofilm in anode, connected to high O2 cathode (oxic) to generate electricity
Ex. Mudwatts
62
Why does anaerobic respiration produce less ATP than aerobic?
Because O2 has the greatest reduction potential (E0) of all electron acceptors
63
Denitirification
Uses nitrate (NO3) from nitrogen fixation as terminal electron acceptor, reduced to nitrogen gas (N2)
Ex. *Paracoccus denitrificans* (facultative anaerobe in soil, depletes soil N, lower crop yield)
Ex. *Escherichia coli* (facultative anaerobe, nitrate reduced to nitrite (basis for nitrite strip test used to diagnose UTIs))
64
Fermentation
Completion of catabolism without the electron transport system and a terminal electron acceptor (process often named after products, ex. lactic, propionic, ethanolic)
* In cytoplasm
* Hydrogen from NADH transferred onto pyruvate
* Generates fermentation products (lactic acid, ethanol), NAD+, ATP (substrate-level phosphorylation)
Also used in clinical identification (ex. *S. aureus*)
65
Domain: *Archaea*, Genus: *Sulfolobus*
Lives in volcanic hot springs, thermoacidophile (high acid and heat tolerance), chemolithotroph
* Oxidizes H2S to H2SO4
* No cell wall, S layer (protein)
66
Chemolithotrophs
Oxidate inorganic sources, transfer electrons to terminal acceptors (usually O2) to generate ATP and PMF via ETC
67
Iron-Oxidizing Bacteria
Oxidize iron compounds as electron source using O2 as electron acceptor
* Generates insoluble ferric hydroxide (toxic)
* Yield little energy due to iron and O2 having a very small E0 difference
Ex. *Acidithiobacillus ferrooxidans*
68
Nitrifying Bacteria
Oxidation of ammonia to nitrate, carried out by 2 genera together:
*Nitrosomonas* - ammonia to nitrate
*Nitrobacter* - nitrite to nitrate
Followed by denitrification
69
Photosynthesis
Two parts:
Light Reactions: Light energy is trapped, converted to chemical
Dark Reactions: Chemical energy used to reduce CO2 and synthesize cell material via Calvin Cycle
70
Oxygenic Phototrophs
Oxidize H2O for electrons, forms oxygen (eukaryotes and cyanobacteria)
71
Anoxygenic Phototrophs
Electrons from other sources (all other bacteria)
72
Chlorophylls
Major light-absorbing pigments in eukaryotes and cyanobacteria (utilize thylakoids)
**Cyanobacteria are Gram -!**
73
Bacteriochlorophylls
Major light-absorbing pigments in purple and green bacteria
74
Accessory Pigments
Transfer light energy of broader wavelengths to chlorophylls, quench toxic forms of oxygen (photoprotection antioxidants), integrated in photosystems within thylakoids (PSI and PSII)
Ex. Carotenoids, phycobiliproteins
75
Reaction Center Chlorophyll
Energy is transferred here after the photosystem absorbs light energy, electrons are excited and passed to the first acceptor in the ETC
76
Light Reactions: Plants and Cyanobacteria
Occurs within thylakoid membranes (chlorophyll)
* Oxygenic (in plants and many bacteria), H2O serves as electron donor/acceptor
* Two photosystems
* Cyclic and Noncyclic photophosphorylation
77
Cyclic Photophosphorylation
Uses PSI, energy from ETC generates PMF to make ATP via F1F0 ATP synthase
78
Noncyclic Photophosphorylation
Uses PS I and II, electrons used to generate NADPH in PSI (which are used in biosynthesis), replenished by splitting water in PSII
79
Z Scheme
Energy diagram for electron transfer in the light reactions of photosynthesis (P680 (PSI) and P700 (PSII))
80
Light Reactions: Green and Purple Bacteria
Occurs in plasma membrane using bacteriochlorophyll
* Anoxygenic (use H2, H2S, or organic matter as electron donors)
* Only PSI (only cyclic photophosphorylation, generates ATP but no NADPH)
81
Photosynthetic Archaea
Utilize **rhodopsin** pigment protein (also found in retina) instead of chlorophyll
82
Microbial Rhodopsin
Pigment protein found in plasma membrane
* 7 Transmembrane helices
* Uses retinal pigment protein to absorb light
* Pumps proton out (light-driven proton pump, rather than ETC)
83
Calvin Cycle
Anabolic pathway for fixing CO2 into carbohydrate, occurs in carboxysomes
* Dark reactions of photosynthesis
* Energy demanding (18 ATP to make glucose)
* Occurs in chloroplasts in plants, cytoplasm in bacteria
* **Crucial to life**: provides organic matter for heterotrophs (use reduced organic molecules as C sources)
84
Many biosynthetic pathways are ____ of catabolic pathways
reversals
85
Gluconeogenesis
Glucose synthesis (aerobic reversal of glycolysis)
* Animals, plants, fungi (humans use to maintain blood glucose levels)
* Requires ATP and GTP
* Uses 6 enzymes from glycolysis, 4 unique enzymes
86
Griffith's Transformation Experiments
Through testing on mice, Griffith found that DNA of dead pathogenic bacteria (*Streptococcus pneumoniae*) was taken up by live non-pathogenic cells, which gained pathogenicity via HGT
87
Gene
Functional unit of genetic information, made of DNA
* Gene notation: *pilA*, *lacZ*
* Protein notation: *PilA*, *LacZ*
88
Genome
All genetic material within a cell or virus
* Consist of usually one, sometimes more DNA chromosomes in bacteria
89
Many genes in bacteria are organized as ___
Operons
90
Operon
A cluster of genes controlled from common regulatory elements
91
Repressor Proteins
Bind to block transcription of operons
92
Inducer Proteins
Molecules that bind to repressors, preventing them from binding to operators, facilitate transcription
93
DNA Structure
Made up of nucleotides containing a pentose sugar (deoxyribose), a nitrogenous base (A/T/G/C), and a phosphate group
Each strand is composed of phosphodiester bonds, they bind to each other using H bonds
* Antiparallel structure of complementary strands, moving 5' to 3' in double helix
94
Purines
Adenine and Guanine
"pure as silver (Ag)"
95
Pyrimidines
Cytosine and Thymine
96
Base pairing rules:
A with T (via 2 H bonds)
G with C (via 3 H bonds)
97
5' GAATTC 3'
3' CTTAAG 5'
Palindrome, restriction enzyme EcoR1 cuts here
98
DNA Size
Expressed as base pairs (bp), can be kbp or Mbp
99
Smallest bacterial genome with only 580 kb encoding 480 proteins:
*Mycoplasma*
100
Prokaryotic DNA
* Usually circular, closed supercoiled molecule
* Bacteria pack DNA into loops or domains, collectively the **nucleoid**
101
Eukaryotic DNA
* Linear
* Eukaryotes wrap DNA around **histone** proteins, collectively called nucleosomes "beads on a string"
* Genes interrupted by introns
Archaeal chromosomes are circular with histones
102
Semiconservative DNA Replication
Strands separate first, each serves as a template strand
103
DNA replication in eukaryotes is ___
bidirectional, with multiple origins of replication (*ori*)
104
DNA replication in prokaryotes is ___
bidirectional, with a single *ori* and two forks moving in opposite directions, ending in termination region (*ter*)
104
How is the prokaryotic ori selected?
DnaA protein binds to distinct sequences within the *ori*
105
DNA Polymerase
Catalyzes DNA synthesis in 5' to 3' direction
* Needs: template, dNTPs, and a primer (RNA with 3' OH group)
* Most bacterial have several, with DNA Pol III being the major replication enzyme
106
DNA Gyrase (topoisomerase)
Underwinds DNA, cuts one DNA and passes another through the gap and seals
* Target for Ciprofloxacin (quinolone) antibiotic
107
DNA **H**elicase
**H**ydrogen bond breakers
108
DNA Polymerase I removes ___
RNA primers and fills in
109
DNA primase lays down ___
RNA primers
110
Single-Stranded Binding proteins (SSBs)
Coat newly separated strands to prevent them from rejoining
111
DNA Ligase forms ___
phosphodiester bonds to seal daughter strands
112
Transcription in Bacteria
* DNA to RNA via RNA polymerase
* Generates 3 RNA types (tRNA, mRNA, and rRNA)
113
RNA Polymerase
* Opens and unwinds DNA without a primer
* Made up of a core and sigma factors
* Targeted by antibiotic rifampin
114
Sigma Factors
Proteins that direct the core of RNA Polymerase to promoter regions and attach
Ex. *o*70, *o*S
115
RNA Structure
* Usually single stranded
* Ribose instead of deoxyribose
* Uracil replaces Thymine
116
tRNA ("transfer RNA")
Converts the language of RNA into that of proteins
* Clover leaf shape
* Two functional regions (anticodon - complementary to codon in mRNA, 3' end - synthetase enzyme attaches an amino acid to it)
* Carries amino acids during protein synthesis (translation) to the ribosome
117
rRNA ("ribosomal RNA") component of ___
ribosomes
118
mRNA ("messenger RNA") is a template for ___
protein synthesis
119
Rho (*p*) Independent Termination
* DNA sequences
* Encodes RNA stem loop structure
* Causes RNA Polymerase to release from DNA
120
Rho (*p*) Dependent Termination
* DNA sequence
* Causes RNA Polymerase to slow due to high levels of G-C 3H bonds
* Protein Rho (*p*) then binds to RNA, catches up to RNA Polymerase and causes release
121
Bacteria use ____ signal transduction systems to control gene transcription in response to their environments
two-component
(also known as Histidine-Kinases)
122
EnvZ/OmpR system senses and controls...
EnvZ **senses high osmolarity**, phosphorylates OmpR response regulator to regulate transcription of structural genes OmpF and OmpC **controlling aquaporin formation** in **E. coli**
123
Transcription in Eukarya
Occurs in nucleus, uses 3 RNA Polymerases
* Has transcription factors (no sigmas)
* TATA box - promoter element
* RNA splicing to remove introns
* mRNA is modified to add 5' cap and 3' poly A tail to stabilize (infleunza virus carries out "cap snatching")
124
Translation is the synthesis of ____ directed by mRNA sequence, requires ____ and ____
polypeptides, ATP, GTP
125
Bacterial Ribosome
The site of protein synthesis, "read" mRNA sequence as a code (codons) and match to corresponding amino acids
* Consists of the E (exit), P (peptidyl), and A (aminoacyl or acceptor) regions
2 subunits: 30S and 50S
* 30S: 21 proteins and 16S rRNA
* 50S: 34 proteins and 23S and 5S rRNA
126
23S rRNA
Peptidyl transferase (**ribozyme**) - catalyzes the formation of peptide bonds between amino acids during translation
127
16S rRNA
Aligns mRNA with ribosome, has sequence complementary to Shine-Dalgarno sequence of the mRNA
128
5S rRNA
Functions in ribosome stability
129
Codon
A nucleotide triplet encoding a specific amino acid or function
* 64 codons
* 61 specify amino acids ("sense")
* 3 are stop codons ("nonsense"), UAA, UAG, UGA
* AUG is the start codon (codes for Methionine)
130
mRNA code is ____, multiple codons can encode the same amino acid
degenerate
131
Translation Phases
1. Initiation
2. Elongation
3. Termination
132
Shine-Dalgarno Sequence
Ribosomal binding site in bacterial mRNA, located 8 bases upstream of the AUG start codon
133
Translation: Initiation
1. 16S rRNA in the ribosome hybridizes with the Shine-Dalgarno sequence of mRNA, which aligns with the ribosome
2. tRNA with formylmethionine binds start codon in P site (anticodons matching mRNA sequence)
134
Translation: Initiation
1. 16S rRNA in the ribosome hybridizes with the Shine-Dalgarno sequence of mRNA, which aligns with the ribosome
2. tRNA with formylmethionine binds start codon in P site (anticodons matching mRNA sequence)
135
Translation: Elongation
1. tRNA binds at A site (requires GTP)
2. Peptide bond joins amino acids, catalyzed by 23S rRNA ribozyme
3. Ribosome moves 1 codon along mRNA
4. Empty tRNA moves from P to E region
Antibiotic tetracycline inhibits binding of tRNA to ribosome complex
136
Translation: Termination
1. Any one of 3 stop codons (UAA, UAG, and UGA), no tRNAs
2. Release facotrs (RF) cleave, release peptide
137
Mutation
* Occurs in all domains
* Heritable change in DNA sequence
* Can generate alleles, give rise to new phenotypes
138
Vertical Gene Transfer (Eukarya)
* Sexual reproduction
* New combinations of genes when gametes of parents fuse
139
Horizontal Gene Transfer (HGT)
Transfer from one independent organism to another via 3 mechanisms:
1. Transduction
2. Conjugation
3. Transformation
140
Transduction
Bacterial gene transfer by phages
Two forms:
1. Generalized Transduction
2. Specialized Transduction
141
Phages
Viruses that infect bacteria
* Abundant and diverse, impact composition and behavior of microbial communities
Two phage types:
1. Virulent Phages - lytic cycles (cell lysis)
2. Temperate Phages - lysogenic cycles
142
Generalized Transduction
* Occurs during lytic cycle, any part of the bacterial genome can be transferred
* During viral assembly, pieces of degraded host DNA can be mistakenly packaged into phage
143
Specialized Transduction
* During lysogenic cycle, a specific part of host genome is transferred
* Prophage incorrectly excises, takes part of genome with it (adjacent to site of integration)
144
Prophage
Sequence of viral DNA that integrates into the host cell during the lysogenic cycle
145
Lysogenic Conversion
* Phenotypic change in a host by temperate phages
Ex. Diphtheria and Cholera toxin genes are encoded by prophages (cell without phage DNA **are not pathogenic**)
146
Phage Therapy
Therapeutic use of bacteriophages to treat pathogenic bacterial infections
147
Conjugation
DNA transfer by direct cell contact
* Requires sex pili and plasmids
* Major mode of spreading antibiotic resistance genes (R plasmid transfer)
148
Plasmids
Double-stranded, circular DNA
* Extrachromosomal (**episomes**), can be transferred by conjugation
* Carry genes that confer advantages
* Are replicons (have own *ori*)
Ex. F Factor in *E. coli* is a well-studied conjugative plasmid (most genes have tra label which are required to transfer itself)
149
Conjugation: F+ x F- Mating
1. Begins with the F+ donor extending pilus to F- recipient
2. Pilus retracts once connection is made
3. Plasmid-encoded enzyme nicks one strand of F factor, allowing DNA Pol III to replicate new strand as single strand enters recipient
4. A new complementary strand is made in the recipient, and in the donor (via Rolling Circle Replication)
150
High Frequency of Recombination (Hfr) Cell
Can transfer integrated F factor and prt of chromosome to F- cell, which becomes an F' cell (**not +**)
