Exam 1 Flashcards
Components of Prokaryotic Cell
Envelope, cytoplasm, nucleoid
Components of Eukaryotic Cell
Chloroplast, cilium, endoplasmic reticulum, golgi, lysosome
Prokaryotic Cell Size
0.1 um - 0.5 mm (5000 fold range) length, most are only 1-2 um
500 fold range width
Eukaryotic Cell Size
10-100 um in length, 2-200 um in diameter
Surface-to-volume ratio
3/r
G+
Thick peptidoglycan layer
Teichoic and lipoteichoic acids provide stability
G-
Lipopolysaccharide layer, thin peptidoglycan
G- Cell Wall is made up of
Outer membrane, peptidoglycan layer
G- periplasmic space
Between cell wall and inner plasma membrane
Peptidoglycan components
Polysaccharide chains of NAG and NAM connected through B 1,4 linkages
Chains connected by short tetrapeptides of repeating amino acid enantiomers (L-ala, DAP, D-glutamic acid, D-alanine)
Regulation of Cell Wall
Autolysins: cleave peptide cross links
Transpeptidases: repair breaks and add additional peptidoglycan
Abundance and activity tightly controlled by the cell
Cell Wall Functions
Determines cell shape, resists intracellular pressure
S-layer
Made of repeating protein subunits (often single protein) that form a sheet-like structure
“liquid crystalline array”
Two dimensional
Pores
Fungal cell walls
Chitin, resembles peptidoglycan. Polymer of B-1,4 linked NAG
Cellulose. Polymer of B-1,4 linked glucose
Glucan. Polymer of alpha-1,6 linked glucode
Algal cell walls
Cellulose or pectin (polymer of galacturonic acid) or protein
Mycoplasma, Thermoplasma cell wall
Lacks cell wall, require environment where external solute concentration is similar to intercellular
Avoid turgor pressure
Bacterial and eukaryotic phospholipids
Use ester bond to couple glycerol and fatty acids
Archaeal phospholipis
Use an ether bond to couple glycerol and fatty acids
Hyperthermophilic archaea unique membrane
Lipid monolayer
Outer membrane is found in _____ and result in the ______
Gram - and gram variable, periplasm
Periplasm
More dilute than cytoplasm
Oxidized
Rapid exchange with environment
Contains hydrolytic enzymes (increases access to substrate, but retains them in close proximity to the cell)
What is the behavioral consequence of the periplasm?
G- secrete less protein into the environment than G+
Lipoprotein
Anchors outer membrane to the cell wall
Tail embedded in inner leaf of outer membrane
Protein component binds cell wall
LPS
Endotoxin, elicits fever in humans
Made of Lipid A, core polysaccharide and O-polysaccharide
O-polysaccharide
made of hexoses in repeating clusters (glucose, galactose, rhamnose and mannose), varies significantly between strains of bacteria. Important distinguishing marker
Core polysaccharide
Varies in composition between organisms
Lipid A
Lipid portion of LPS, ester-linked FAs coupled to a disaccharide of N-acetyl glucosamine phosphate
Gram Variable
Acid fast
Mycobacteria
Made of phospholipid combined with mycolic acids, not LPS
Porins present
Mycolic acid
Long chain fatty acids extending 70 or more carbons
Small molecule transport mechanisms
Simple diffusion, osmosis, facilitated diffusion and active transport
Group Translocation is found in
Prokaryotes, chemically modifies incoming molecule consuming energy
What type of transport is only found in eukaryotes?
Pinocytosis and endocytosis
Cytoplasm
90% water
Highly reducing
Protein thiol groups remain protonated, minimizing sulfhydryl bonds
pH is constant, near neutral
Prokaryotic cytoplasm
Not disorganized
Cryoelectron microscopy and biochemical/genetic analysis of protein complexes show that cellular processes occur in discrete locations, and involve the coordinated action of large complexes of proteins
What grows at pH
Archaea and possibly a few fungi
What grows at pH >10?
Bacteria and archaea
Eukaryotic cytoplasm
Complex
Contains nucleus and organelles
Facilitate movement of proteins and other molecules
Cytoskeleton
Cytoskeleton
Critical for cell division and movement
Crenarchaeal chromosomal organization
NAP (bacteria like)
Euryarchaeal chromosomal organization
Eukaryote like, histone proteins and nucleosomes
Primary transcript processing in Eukaryotes
Splice out introns, 5’-methylguanosine cap, 3’ polyadenylate tail
Prokaryotic ribosomal subunits
30S + 50S = 70S
Eukaryotic ribosomal subunits
40S + 60S = 80S
Prokaryotic 30S subunit
16S rRNA
Prokaryotic 50S subunit
23S and 5S rRNA
Proteasome
Multisubunit macromolecular structures that degrade proteins
Barrel shaped, central pore through which protein substrates pass during degradation
20-26S
Archaea (fewer proteins but higher copy number) and Eukaryotes (25 proteins)
Storage Bodies
Polyhydroxybutyrate inclusions
Carboxyl connected to carbon chain, R group determines type > CH3 = PHB
Nitrogen rich compounds
Lipids
Capsules
Outside prokaryotic envelope Glycocalyx or slime layer Polysaccharides (or amino acids) that coat cells (type varies) Escape from predation Surface adherence Virulence (escape phagocytosis) Resistance to environmental extremes Loss can lead to nonpathogenicity
Pilus
Also called fimbria
Made of pilins arranged in a helical array forming a long thin tube-like structure with a hollow central core
Interspersed with adhesins (enable attachment)
G- sex pilus allows conjugation
Loss can lead to nonpathogenicity
Flagellar Motor
Nanomachine
Rotate flagellum around axis
Made of Mot, Fli proteins, M and S rings, P and L rings
Flagellin
Protein subunits that make up flagellum
Bacterial vs Archaeal Flagella
Flagellin is similar among bacteria, but different in Archaea
Flagella evolved separately in the two groups
Flagellin is assembled at
The tip of the rod, not the base
Flagellin monomers pass through the hollow core where they are assembled by cap proteins
Assembly of flagellum
Insertion of M and S rings, motor proteins, then P and L rings and finally the hook
Mot
Rotation of the motor is caused by mot (motor) proteins
Use proton gradient
Size of the gradient is proportional to rate of rod movement
1,000 proton required for each rotation
Fli
Control direction of rotation, reversible depending on intracellular signals concerned with environmental conditions
Oligotrophy
Prefer conditions of limited nutrient availability
Do not exhibit unrestricted growth under nutrient excess conditions, divide at very slow rates
Open ocean
Copiotrophy
Exhibit unrestricted growth in response to conditions of nutrient excess
Model systems, achieve high densities in laboratory culture
Colony with diameter of 1 mm
10^8 cells (100 million)
Spectrophotometry can detect
Populations 10^7 cells or greater
Changes within a colony
Differences in oxygen, light availability, pH, nutrients and osmolarity
Death Phase
Not obligatory, not terminal step in pathway of differentiation
Microbes can stay in growth phase indefinitely
Stationary cells can transition to lag and then growth phase to avoid death phase
Quorum Sensing
Signal transduction
Environmental or biological cue that is communicated to cells and stimulates changes in gene expression
Bacterial prokaryotes and fungal eukaryotes
Can results in macroscopic events
Quorum sensing molecules
Homoserine lactones
Small peptides
Flourescence
Luciferase encoded by lux genes
Oxidizes long chain aliphatic aldehydes and flavin mononucleotide in the presence of oxygen
Products are light, water, oxidized flavin mononucleotide and long chain aliphatic carboxylic acid
Tooth biofilms
Streptococcus mutans and Fusobacteria
Consortia
Simple mixtures of taxa that collectively conduct biochemical reactions that convert a starting substance into one of applied or environmental importance
“biotransformations”
k
Relationship between N and t
Specific growth rate constant
k=
(logN2-logN1)2.303/(t2-t1)
in hrs^-1
g
generation time
dependent on k
g=
ln2/k
in hrs
Balanced Growth
When oligotrophic microbes are supplied with unlimited food and permissive growth conditions, they grow and divide at maximum rates
All components of a population of cells and single cells increase proportionally
What is used to measure RNA synthesis?
Radioactive uracil
Submaximal growth rates
Restricted by nutrient availability or environmental conditions
Why microbes are present in most natural samples at lower densities
Cell Yield
Absolute number of cells that can be produced
Determined by nutrient availability and the type of metabolism
Cell Yield Copiotrophs
Directly related to the limiting nutrient available or type of metabolism
Cell yield is half when the amount of the limiting nutrient is halved
Cell Yield Oligotrophs
Relationship with nutrient availability not clear
More restricted metabolisms
Lithoautotrophs seldom achieve cell densities greater than 10^6/mL
Secondary metabolism
Unique metabolic processes during stationary phase
Ex: synthesis of antibiotics and pigments, use of alternate energy yielding and consuming pathways
Richard Morita
Starvation induct stationary phase plays critical role in survival of microbes in the ocean
Stress Resistance Response
Evolutionarily conserved
Hsp70 (DnaK) protein chaperone present in all bacteria and some Archaea
Widely distributed among eukaryotic microbes in mitochondria and chloroplasts
Hsp70
Eukaryotes
DnaK
Prokaryotes
dnaK mutant
Only modestly sensitive to heat during growth but extremely sensitive during stationary
Protein refolding is critical
Size and stationary phase
Cells become smaller
Differences are taxon-specific
Reductive division
Changes shape and increase surface are to volume ratio, increases ability to scavenge nutrients
Viable but notculturable cells (VBNC)
Reversible, obscures the criteria for viability
Vibro cholera
Rita R Colwell
VBNC vibrio cholera research
How can viability be shown in VBNC cells?
Nutrient dependent increases in cell size
Enterobacteria
VBNC, present in chlorinated drinking water and survive disinfection
Low level residuals must be maintained to maintain bacterial injury
Maintenance energy
Energy consumed for cell maintenance
Transporting nutrients and waste products across envelope
Intracellular detoxification
Motility
Protein refolding
Becomes a large fraction in stationary phase
Energy storage compounds
Stationary Genetics
Disable genes for DNA repair
Increase in spontaneous mutation
Mutators defective in mismatch repair pathway
Proteins naturally become depleted in stationary, elevates homologous recombination and mutation rate
Layers of the Spore
Core, cortex, coat, exosporium
Endospore coat
Comprised of multiple layers of specific proteins
Deposited in a particular order but not in synthesized order
Spore coat involved a process of auto assembly mediated by the proteins themselves
Dipocolinic Acid
Diagnostic marker of endospores
Cortex
Overlapping mechanisms to confer thermal tolerance
Core
Compressed version of a vegetative cell with DNA contained within a cytoplasmic space and a membrane
Cortex
Surrounds the core and consists of peptidoglycan
Protein Coat
Made up of multiple layers of spore specific proteins
Exosporium
Surrounds the cortex and consists of a unique but thin layer of special protein
Sporulation Stages
1: Vegetative cell
2: Septation and engulfment
3: Prespore protoplast formation
4: Cortex formation
5: Coat formation
6: Maturation
7: Spore release
spo genes
Control sporulation
Mutations in any result in defective sporulation
spo mutants elucidated steps of sporulation
Spo0A
key regulatory protein that initiates sporulation
phosphorylated or dephosphorylated
Sporulation sigma factors
sigmaA: vegetative sigma
sigmaE: prepares parental cell, directs expression of genes specific to vegetative cell surrounding the endospore
sigmaF: prepares parental cell, directs gene specific to the endospore
sigmaG: activate genes necessary for terminal stages of endospore development
sigmaK: gene expression in the surrounding vegetative cell late in development
sigmaF activates
sigmaG, activates genes for terminal endospore development
sigmaE activates
sigmaK, activates genes for terminal development in the vegetative cell
Biofilm Stages
- Transient attachment
- Permanent attachment
- Maturation
- Detachment
EPS
extracellular polymeric substances
excreted slime layer
Channels and pores permeate during maturation, increase supply of oxygen and other nutrients
Provide structural integrity
Transition to permanent attachment
Cell synthesizes EPS
OR
Production of specific proteins including pili, fimbriae, curli and adhesins that promote specific attachment
Signal molecules biofilms
Homoserine lactones in proteobacteria
Peptides in Bacillus and Streptococcus
Endocarditis
Infection of cardiac valve
Vegetation: bacterial cells combined with platelets and fibrin
Compromises blood flow and valve function
Recurrent source of infection
Cause of embolizatoin
Prolonged antibiotic therapy and removal of valve
Cystic Fibrosis
Congenital disease Lung biofilms Environment caused by mutations in CFTR promotes colonization and biofilm development Pseudomonas aeruginosa Death by asphyxiation
Kidney Stones
15-20% accompany UTIs and derive from biofilms
Biofilm combine with struvite (magnesium phosphate and calcium apatite)
Forms when urease producing organisms create alkaline environment
Biofilms and Evolution
Density is higher than planktonic
Increased rates of horizontal gene transfer
Evolution of pathogens
Functions of Transport Systems
- Concentrative
- Discriminating
- Regulatory
- Versatile
Importance of Transport Systems
- Efflux system
- Protectant uptake
- Environment sensing
- Catabolite repression
- Flagellar motor
- Ion regulation
- Proton translocation
- Nutrient acquisition
Percentage of gene products that are transmembrane
18-20%
Milton Saier
Transportation Classification System
Channels/Pores Example
Glycerol permease
Glucose permease in Zymomonas mobilis
Entner-Doudoroff Pathway
1 glucose > 2 ethanol + 2 CO2 + 1 ATP
Yields less ATP than normal glycolysis
Primary Active Transport
F0F1 ATPase
Kdp system
Na+ transporting carboxylic acid decarboxylase
Kdp system
Pumps K+ into the cell using ATP
Na+ transporting carboxylic acid decarboxylase
Creates Na+ gradient to bring citrate into the cell
Uses energy from decarboxylation oxaloactetate to pump Na+ back out
citrate > oxaloacetate + acetate > pyruvate + CO2
ABC systems
ATP Binding Cassette High substrate affinities Substrate binding protein Integral membrane spanning proteins Inner membrane associated ATP hydrolysis proteins
Ionophores
Dissipate ion gradients and inhibit secondary transport
Osmotic Shock
Expose cells to in hypertonic buffer to EDTA
Removes divalent cations that hold together LPS
How do anaerobes makes a pmf?
F0F1 ATPase
How to measure transport and identify energy source?
- Radio-labeled substrates
- Control source of energy
- Use vesicles or proteoliposomes
- Mutational analysis
Product:precursor exchange systems
Oxalate > formate Arginine > ornithine Lactose > galactose Citrate > lactate Malate > Lactate (malate permease)
Group Translocation
PEP dependent PTS
PEP > pyruvates phosphorylates Enz I which phosphorylates Hpr which phosphorylates Enz II which is substrate specific
Hpr and Enz I
Cytoplasmic and not substrate specific
Enz II
Multiple domains One integral (Enz IIC) Substrate specific
Hpr
Key intermediate phosphorylated at His15l
Acid tolerance
HA is pumped in/out of the cell were it can dissociate
HA= acetic, propionic, benzoic acid
Acid tolerance
Pump H+ out of cell
Pump malate in, convert to lactate using H+ and producing CO2, lactate leaves
Pump arginine in which creates ammonia, CO2, ornithine (pumped out)
Osmotolerance
Betaine is pumped out or into the cell
Cryotolerance in Saccharomyces
Trehalose permease symports trehalose with H+, can produce glucose as intermediate
