Lecture 1 - Exam 1 Flashcards
How much do we know about microbes?
Considering their importance, it is surprising how little we know about their workings.
Why study microbial physiology?
Microbes play a central role in our existence. They help us grow, preserve, and digest our food. They can spoil food and poison us. They produce life-saving antibiotics/antifungals and life-threatening infections. They play key roles in nature by helping global recycling of nutrients and energy (they fix nitrogen in nitrogen cycle).
What are the different types of prokaryotes?
Halophilic, psychrophilic, thermophilic, cannibalistic.
What is a halophile?
Can be both archaea and bacteria. Can be found in the dead sea, live in very salty conditions.
What is a psychrophile?
In permafrost and arctic waters, can be recovered from glaciers. Cold-loving prokaryotes. Bacillus anthracis spores recovered in Russian permafrost from 13th century.
What is a thermophile?
Live in hot springs and geysers. Hot-loving prokaryotes. Responsible for the beautiful colors of Grand Prismatic Spring.
What is a cannibalistic prokaryote?
Vampirococcus lugosii is a cannibalistic prokaryote with a small genome and limited biosynthetic metabolic capabilities. It lives in anaerobic, aqueous conditions. They perforate the cell wall and membrane of their victim Chromatiaceae (purple sulfur bacteria) and sucks out its cytoplasmic contents.
What is the nomenclature for genes?
3 lower-case italicized letters followed by an upper-case letter that differentiates alleles.
Example: rpoB (italicized though)
What is the nomenclature for proteins?
They are not italicized, and the first letter is upper-case.
Example: RpoB
How do you write out the genus and species of organisms?
Genus is italicized and the first letter is upper-case. Species is italicized and lower-case. You should write out the full genus and species the first time, and afterwards, the genus is abbreviated.
Example (First time): Bacillus anthracis (italicized)
Example (After the first time): B. anthracis (italicized)
What are the three domains of life? How many phyla in each domain?
Bacteria: >40 phyla
Archaea: >20 phyla
Eukarya
The three domains of life are based on what?
16S Ribosomal RNA gene sequences.
What is our model organism in the Bacteria domain?
Escherichia coli
Gram-negative bacterium
Proteobacteria
LUCA stands for what?
Last universal common ancestor
What are the methods we use to study prokaryotic physiology?
Microbial techniques: growth rates, nutritional requirements, temp. range, pH range, O2 requirements, cellular interactions etc.
Microscopy: light, fluorescent, electron, confocal, atomic force
Genetics: isolation and characterization of mutants, study of gene expression, genetic manipulation
Molecular biology and biochem: characterize genes and proteins that cause the mutant phenotypes, examine separate parts and functions in vitro, “omics,” systems biology, synthetic biology etc.
We combine these various approaches to piece together the puzzle.
How many prokaryotes are there? Are most prokaryotes unculturable?
There are an estimated 2.2-4.3 million species. Most prokaryotes are uncultured (we just don’t know quite how to do it yet), not that they are “unculturable.”
What is the endosymbiotic theory?
That the origin of the first Eukaryotic cell was from an Archaea (within the Asgard superphylum) that engulfed an alpha-Proteobacteria.
What are the characteristics of a prokaryote?
No membrane-enclosed organelles: no nucleus, mitochondria, golgi-complex, endoplasmic reticulum, etc.
However, they have many internal structure: thylakoid membranes for photosynthesis, carboxysomes for CO2 fixation, etc.
What does the organismal diversity look like for prokaryotes?
Largest biomass of living organisms, nutritional versatility (can digest carbs, hydrocarbons, organic acids, almost anything), metabolic diversity (nitrogen fixation, sulfur reduction/oxidation, photosynthesis, etc.), modern phylogeny determined by 16S rRNA gene sequence.
Information processing (DNA -> RNA -> Protein) is very similar in…?
Archaea and Eukaryotes. This is not surprising due to the endosymbiotic theory.
What is RNA polymerase (RNAP)? Who uses it? How many do they use?
RNAP is an enzymes used to copy DNA sequences into RNA sequences. Archaea, Bacteria, and Eukaryotes use RNA polymerase, but both Archaea and Bacteria use only 1 type of RNAP to transcribe all genes, whereas Eukaryotes use 4 RNAPs (I, II, III, IV). Archaeal RNAP is much more complex than Bacterial RNAP.
Describe Bacterial RNA Polymerase.
Bacteria RNAP has 6 subunits (Rif^S). Bacterial RNAP is Rifampcin-sensitive (Rif^S), meaning Rifampcin inhibits bacterial RNAP (binds with high affinity).
Describe Archaeal RNA Polymerase.
Archaeal RNAP has 12-13 subunits. Archaeal RNAP has similar structure to eukaryote RNAP II. Both Archaeal and Eukaryotic RNAP is resistant to Rifampicin (Rif^R).
What are the histones like in Archaea?
Archaea contain eukaryotic-like histone proteins that compact archaeal DNA into compact structures.
What are the ribosomes like in Archaea?
Archaeal ribosomes are similar to that of eukaryotes. The ribosomes are sensitive to Diphtheria toxin, like eukaryotic ribosomes.
What does the Diphtheria toxin (that archaeal and eukaryotic ribosomes are sensitive to) do?
The Diphtheria toxin inactivates elongation factor (EF2), inhibiting protein synthesis.
What is the structural and functional difference between Archaea, Bacteria, and Eukarya? (8 characteristics to cover)
{Peptidoglycan:
Bacteria - yes ; Archaea - no ; Eukarya - no
{Lipids:
B - ester linked ; A - ether linked ; E - ester linked
{Ribosomes:
B - 70S ; A - 70S ; E - 80S
{Initiator tRNA:
B - Formylmethionine ; A - Methionine ; E - Methionine
{Introns in tRNA:
B - No ; A - Yes ; E - Yes
{Ribosomes sensitive to Diphtheria Toxin:
B - No ; A - Yes ; E - Yes
{RNA polymerase:
B - One (5 subunits) ; A - One (12-13 subunits) similar to eukaryotic RNAP II ; E - Four (12-14 subunits each)
{Ribosomes sensitive to Chloramphenicol, streptomycin, and kanamycin:
B - Yes ; A - No ; E - No
Moving from the outside of the cell to the inside of the cell, what are the major cell components (order is important here)?
- Appendages (flagella and pili)
- The Glycocalyx
- Cell Wall
- Cell Membrane
- Cytoplasm
What are the cell structure differences between Archaea and Bacteria?
Cell walls, cell membranes, and flagella
What are bacterium flagella?
Flagella are tail-like structures primarily used for motility in aqueous habitats. They also may be involved in adhesion and host-cell invasion by some bacteria.
Flagellar synthesis is a highly regulated process.
What are the different numbers and arrangements of flagella?
Monotrichous = 1 flagellum
Peritrichous = all over the surface
Lophotrichous = in clumps (bundled)
Amphitrichous = one flagellum at both ends
What are the three basic parts of a flagellum?
Filament, hook, and basal body
What are the characteristics of the filament of a flagellum?
Extends to the exterior and is made of proteins called flagellin (that are added at the tip).
What are the characteristics of the hook of a flagellum?
Connects filament to the cell.
What are the characteristics of the basal body of a flagellum?
Anchors flagellum into cell wall.
Flagella also have what embedded into the cell wall? What do these things do?
Flagella have several protein rings embedded in the cell wall that act as molecular motors and are powered by ATP.
Is the production of flagella energetically-unexpensive or energetically-expensive? Example?
Energetically-expensive.
Flagellated forms of bacteria grow 2% more slowly than their non-motile forms.
The expression of motility genes is regulated at which levels?
The expression of motility genes is regulated at practically all levels: Transcription, translation, and transport
In bacteria, flagellar growth occurs where?
In bacteria, growth occurs at the distal end (flagellin subunits added to the tip of the flagellum).
Flagellar gene regulation is coupled to what?
Flagellar assembly
What are the classes of temporally-expressed genes?
Early, middle and late-stage expression (Class 1, 2, & 3). One cluster is expressed first, then the expression of the second cluster will turn on, then the third. This is done in this way to save energy.
What are the early genes, and what do they do?
The master operon genes. These are controlled by numerous global regulatory signals. These signals result in the expression or inhibition of flagellar gene expression.
The flagellum and its associated components are assembled in a precise order. What is this order? (Slide #26 to look at diagram)
- MS and C rings (the Rotor) : assembled in a coordinated fashion
- Construction of transport apparatus: exports the flagellar components through a channel in the center of the MS ring.
- Rod and Hook: The hook is not completed until P and L rings are assembled.
- Filament: Flagellin monomer export follows hook completion, and the filament is assembled (in bacteria the filament grow from the tip, but in archaea, subunits are added at the base)
- MotA and MotB (the stator): Assembled concurrently with the filament. Last things expressed, and w/o them then completely unfunctional.
The timing of assembly of the components is (at least partially) controlled at the which level?
The level of transcription for the genes encoding the components.
The flagellum genes are contained in three gene classes (early, middle, and late-stage expression). There are also three temporally-expressed clusters. What are they?
Cluster 1: Master Regulator: FlhDC is the transcriptional activator of Cluster 2 genes. Only class one genes here.
Cluster 2: Contains genes that code for the basal body and hook, as well as the sigma factor (FliA). Also, contains class 2 and class 3 genes. The class 3 genes in this cluster code for the filament and cap.
Cluster 3: Contain genes that code for the stator (the power generator of the motor), MotA and MotB and the anti-sigma factor, FlgM. Only class 3 genes here.
What does the sigma factor FliA (has responsibilities during flagellum assembly and gene expression) do?
FliA initiates transcription of Class 3 genes. Without it, there would be no class 3 genes, meaning the cell would not be able to turn.
What does the motor of the flagellum do, and what are its two parts?
The motor causes the flagellum to rotate and includes the stator and rotor.
Describe the stator in the flagellum.
The stator is one part of the motor. It does not rotate, and is comprised of MotA and MotB proteins.
Describe the MotA and MotB proteins that comprise the stator part of the flagellum.
MotA and MotB form a complex that spans the cell membrane and surrounds the MS ring. The MotA/MotB complex conducts protons (proton motive force) from outside the cell to inside the cell, powering the rotation of the rotor. As the protons move through the complex, this changes the conformation of the complex and affects the rotor.
Describe the rotor in the flagellum.
The rotor is the MS ring and C ring together. Within the C ring, FliG interacts with the Mot proteins, and this transmits the torque required to rotate the flagellum. To control rotation, the regulatory protein CheY (switches flagellar rotation on or off) binds to the C ring.
One of the few known examples of biological rotatory machines, the _______ is unique in its remarkable power and efficiency in converting free energy into mechanical work.
Motor
Describe flagella-mediated locomotion.
Flagellar motor of plain filaments turns counterclockwise by default. This allows several peritrichous flagella to bundle together and propel the cell smoothly forward. Complex filaments can only rotate clockwise (less common, ex: Pseudomonas & Rhizobium).
In response to the chemotaxis signaling molecule, phospho-Chey, the motors can switch to clockwise rotation. This causes dissociation of the peritrichous flagellar bundle and results in a tumble. Allows for reorientation of the cell.
What is Tactic Response of E. Coli?
It is the ability to move in response to environmental stimuli.
The cell has sensory control of swimming behavior and environmental change is sensed by a sensory protein.
What type of protein is responsible for sensing the external environment, regarding tactic response?
Transmembrane proteins, and some species may have additional cytoplasmic receptors.
What are the types of tactic responses?
Chemotaxis: Chemical stimuli
Phototaxis: Light stimulus
Aerotaxis: Oxygen
Magnetotaxis: Magnetic Field
What are the two important considerations when we talk about tactic response?
- Length of run
- Direction of run
Bacteria measure the chemoeffector over time. How does an attractant and repellent effect the bacteria?
If bacteria sense an attractant, tumble frequency decreases (runs are longer).
If bacteria sense a repellent, tumble frequency increases (runs are shorter).
What is swarming motility?
A form of social swimming in which cells move on a wet, solid surface. Bacterial populations that swarm can rapidly spread as a multicellular population rather than as single cells.
What does swarming motility do to new cells? How about competition for nutrients?
Will direct new cells to the edge of the colony. It reduces competition for nutrients between cells, speeding growth.
How do the cells in a swarming population change their morphology in liquid?
Swarmer cells become more filamentous, and can have many lateral flagella. They use the lateral flagella to physically interact with other cells in the swarming population.
What type of cell has internal flagella?
Spirochetes
Where are the internal flagella located in spirochetes? What do these internal flagella do?
Located in the periplasm between inner and outer membrane - “periplasmic flagella.” The internal flagella wrap around the length of the cell and overlap in the middle. The rotation of axial filaments creates “cork-screw” movement.
What is so special about the cork-screw movement of spirochetes?
Swimming using the cork screw movement allows the pathogen to be highly invasive and virulent in human hosts. Ex: Treponema pallidum
What do Type III Secretion Systems (T3SS) do? Where could it of arisen from?
Uses a flagella-like needle (injectisome) to inject effector proteins into eukaryotic target cells. T3SS may have arisen from exaptation of the flagellum (i.e. the recruitment of part of the flagellum structure for the evolution of the new protein delivery function). So, flagella and T3SS secretion systems are paralogous gene products.
Describe the Archaeal flagellins.
Possess a highly conserved hydrophobic N-terminal sequence that is similar to that of type IV pilins and clearly unlike that of bacterial flaggelins.
Describe the archaeal flagellum: Archaellum.
It is unique to Archaea, in that it is distinct in composition and assembly from bacterial flagellum. There are no homologs of any bacterial flagellar genes that have been identified in Archaea. The archaellum is assembled from the base, not the tip.
Describe Eukaryotic Flagella.
Also function in motility. Completely unique structure unlike that in Archaea or Bacteria.
How many times has flagella evolved?
Bacteria, Archaea, and Eukarya organisms all have completely unique flagellar structures. This means flagella has evolved independently three times. This makes motility extremely important for an organism’s fitness.
Describe gliding motility using a “Tank Tread” like structure.
This kind of motility is on a solid surface, does not use flagella. Ex: Cytophaga use this type of motility. Single-cell gliding is mediated by elastic, viscous, and capillary interactions between the bacteria, the slime it secretes, and the substrate underneath.
