Lecture 8 Flashcards by Andrew Monge

Name the growth phases in order of a population of bacteria inoculated into a flask of fresh media

Lag
Exponential growth
Stationary phase
Death

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Lag phase

No net growth

When we transfer cells from stationary phase of growth to fresh media

The time required for physiological adaption

New enzymes or coenzymes must be synthesized

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Exponential growth

Increase of cell mass exponentially with time

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Stationary phase

No net growth

Exhaustion of nutrients

Limitation of oxygen

Accumulation of toxic growth

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Death phase

Death is greater than growth

Depletion of cellular energy

Activity of autolytic enzymes

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Adaption responses to nutrient limitation in to types of cases?

Lab condition: stationary phase

Natural environment often low in nutrients

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When environments has a low nutrient or is in stationary phase. What are the two types of responses can happen depending on bacteria?

Bacteria 1: sporulate or form cyst to become metabolically inactive or less active

Bacteria 2: Do not form spores But undergo changes which result in resistant metabolically less active cells.
Ex: E.coli, salmonella, vibrio, pseudomonas

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How do bacteria change in cell size due to nutrient limitation?

Bacteria cells use reductive division. Dispersing the population (5-10 micro meter to 1-2 micro meter)

Self digestion of cell material

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Bacteria morphological changes to nutrition limitations?

From rod shape to concord shape

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Changes in surface properties due to nutrient limitation?

Some marine bacteria become more hydrophobic and more adhesive.

Synthesize fibrils, cell aggregate and attach to surfaces.

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List the changes in metabolic activity in nutrient limitation?

Overall metabolic rate slows

Protein and RNA (rRNA) serve as an energy source to maintain crucial cell function

solute transport system
energized membrane
ATP pool level

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Changes in protein Composition

Mays synthesize 50-70 new proteins under condition of carbon, nitrogen, or phosphate starvation:

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Changes in resistance to environmental stress

Cell in stationary phase are more resistant to:

High temp

Osmotic stress

Certain chemicals Like H2O2

Acidic or Basic pH

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Resistant properties are due to?

Synthesis of starvation sigma factors

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What are the names of sigma factors

Rpos (RNA polymerase sigma S)

Sigma S

Sigma 38

Master regulator of stationary phase response

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Sigma factors help RNA polymerase do what?

Sigma factors help RNA polymerase to bind to specific promoter.

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Rpos gene in E.coli encodes?

Sigma factors required for transcription of genes expressed during stationary/starvation phase.

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Protein binds to RNA polymerase and direct transcription of

rpoS- dependent Genes

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What is Rpos required for?

The transcription of a regulan that includes at least 50 genes that encode proteins induced by carbon starvation

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Catalase

Resistance to H2O2

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Exonuclease III

Repair DNA damage

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What is used as a an energy source to maintain crucial cell functions

Protein and RNA (rRNA)

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What are crucial cell functions that are needed to be maintained in stationary phase?

Solute transport systems

An energized membrane

ATP pool level

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In phosphate starvation what is made?

Phone poring in outer membrane to bring in more phosphate

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In nitrogen starvation what is induced?

The induction of N2 fixation genes

What types of starvation can they go under?

Carbon , nitrogen, phosphate starvation

Rpos is what type of regulator?

It is a global regulator

Rpos is important in stress response during what phase?

Exponential growth

Rpos is a stress response to what type of stress?

30 to 40 degrees Celsius High osmolarity

What are persister cells?

A sub population of slow growing or growth arrested bacterial cells that have a decreased susceptibility to antibiotics.

Experiments have shown that persisters cells live in low numbers in this stage?

Late log phase

Experiment have shown that persisters cells increase in numbers in this phase?

Stationary phase

What are the importance of osmotic pressure?

1. Cytoplasm of bacterial cells have more positive osmotic pressure than medium 2. Incoming water expands the cell membrane , which exerts pressure outwards against wall 3. pressure exerted against wall is called turgid pressure

Which component has more positive osmotic pressure the medium or bacterial cell?

The bacterial cell has more positive osmotic pressure.

Incoming water expands the cell membrane. Where does the pressure of the water get exerted on?

The cell wall

Define turgor pressure

Pressure exerted against cell wall

Trugor pressure is necessary for?

Growth of cell wall and cell division

How can you decrease turgor pressure?

Increase osmolarity of medium

What does the increase osmolarity of medium cause?

It causes stop of growth and inhibits variety of physiological activity.

Specify the types physiological activity That is inhibited during increase of osmolarity in medium?

Nutrient uptake & DNA synthesis

During adaption to high-osmolarity. Bacteria increase or decrease intracellular solutes ?

Increase

Intracellular concentrations of certain solutes are called

Osmolytes

Osmolytes are called

compatible solutes

How do bacterial cells get osmolytes?

- some are made by the cells | - some are accumulated via transport

Organic compatible solutes are called?

Osmoprotectants

List 4 types of osmolytes

1. K+ 2. Amino acids glutamate ,glutamine, and prolines 3. Quarternary amine betaken 4. Certain sugars (trehalose)

When E.coli is shifted to high osmolarity

1. Influx of K+ 2. Rapid accumulation of glutamic acid Ionizes K+ 3. Potassium glutamate is replaces with trehalose due to excretion of K+ and glutamate 4. If proline or betain os present in medium this replaces trehalose

Osmolarity effects what type of activity?

Transcription and enzyme activity

The effect of osmolarity causes bacteria to synthesize

New enzymes or transporters. Which are responsible for solutes or transport

Increased transcription is due to?

RNA polymerase sigma S (RpoS)

In S.aureus what is the effect of being shifted to high osmolarity media? transcription and enzyme activity response?

S. Aureus activates pre existing proline uptake system

In E.coli and S. Typhimurium what is the effect of osmolarity on transcription and enzyme activity response?

Increase of transcription of proU, an operon that codes for proline (and betain) transport system

E.coli increase transcription of what in high osmolarity? What does this operon code for?

KDP operon KDP operon codes for high affinity K+ transport system

Bacterial cells must able to adjust to low osmolarity media, to limit?

Turgor pressure

How does the bacterial cell decrease cytoplasmic osmolytes in low osmolarity media?

- excretion | - catabolism

When E.coli is places inside a dilute (hypoosmotic medium) the bacterial cells are?

Hypo-osmotically shocked

Mechanosensitive channels are gated. What cause conformational change?

Water rushing in increasing turgor pressure transiently opening channel alllowing solutes to come in.

How bacteria sense external osmolarity and transmit appropriate signals to genome

Is mostly unknown

The best studied signal system to responding responding in change of external osmolarity include genes for?

OmpF & Ompc Poring in E.coli and salmonella

The total amount of the two proteins are fairly constant when do ratio change?

With osmolarity and temperature

Components Of the structure of trimeric porin

A channel Protein that holds the together And protein that narrows the channels

In high osmolarity transcription of what omp gene?

OmpC to increase ompc channels smaller channels

When e.coli is grown in prescience of 1% NaCl and 30 degree Celsius what type of channel is made? (Simulation of conditions inside intestine)

Only ompc

When E.coli is grown at lower temps and osmolarity. Condition of lake and streams. Which channel is made?

OmpF

(High osmolarity media) What is EnvZ?

A histidine kinase that has been postulated to function as a osmotic sensor

(High Osmolarity media) High osmotic pressure activates or deactivates Kinase activity of EnvZ?

Activates EnvZ

(High osmolarity media) What is a response regulator of EnvZ?

Cytoplasmic protein called OmpR, which receives a phosphorylation group

(High osmolarity media) High level of OmpR-P stimulates transcription of?

OmpC and OmpF by binding to low-affinity sites upstream from their promoters

(High osmolarity media) OmpR-P stipulates transcription of OmpC and OmpF what is the third gene?

The third gene is Mic F from OmpC promoter

(High osmolarity media) Mic F is complementary to?

The 5’ end of ompF mRNA

(High osmolarity media) Mic F is what strand of RNA

Antisense

(High osmolarity) Mic F binds to ompF mRNA where?

Translation and starts and blocks translation

(Low osmotic pressure) Are the concentration of OmR-P high or low?

In low osmotic pressure concentration of OmpR-P are to low

When the concentration of OmpR-P are low it binds to what and why?

It can only bind to high affinity binding site. The binding site is upstream from ompF promoter

(Low osmotic pressure) When OmpR-P binds to bind upstream from ompF promoter results in?

Higher expression of ompF gene

What is stringent response?

The control of ribosomal RNA synthesis.

Ribosome synthesis is coupled to?

Bacterial growth

Amino acid starvation results in?

The inhibition of ribosome synthesis.

A very important regulator of stringent response?

Guanosine tetraphosphate (ppGpp) OR pppGpp

The stringent response is

Temporary inhibition in the synthesis of ribosomal RNA and transfer RNA. When starved by amino acids

In the stringent response. The lack of amino acids causes?

Uncharged tRNAs accumulation, which in turn results in “stalled” ribosomes.

Uncharged tRNAa are also known as?

Non aminoacylated tRNA

The enzyme on stalled ribosomes is called?

RelA (ppGpp synthase I)

Activated RelA synthesizes

The signaling molecule ppGpp and to lesser degree pentaphosphate (pppGpp). Together referred as (p)ppGpp

PpGpp interacts with RNA polymerase and does what?

Down regulates transcription of ribosomal RNA

Advantages of cell- cell communication in bacteria

- To migrate to a more suitable environment/ better nutrient supply - To adopt new mode of growth, sporulation of biofilm formation - Timing of production and release of virulence factors in pathogenic bacteria

autoinducers are molecules that are used for?

Cell- cell communication

Autoinduducers are self generated signal molecules true of false?

True

Autoinducers are used to regulate?

Behavior

Autoinducers regulate behavior of bacteria according to?

Population density (of themself) Other bacteria

Through the use of autoinducers, bacteria can regulate their behavior according to population density of themselves and also bacteria. What is this called?

Quorum sensing

Soluble signals transferred between cells are also called?

Pheromones

Signaling molecules regulate how many functions and what are they?

They regulate 6 functions 1. DNA exchange between bacteria 2. Animal pathogenesis 3. Antibiotic production 4. Bioluminescence 5. Bacteria development 6. Other activity necessary for survival

When autoinducers is release from a single bacterium in a low density colony what’s happens?

Concentration of AHLS are to low to be detected.

When a sufficient density of bacteria are present and autoinducers are released what happens?

Concentration of AI’s reach a threshold that allows the bacteria to sense critical cell mass

What is the response of bacteria when concentrations have reach a threshold?

Bacteria in response activate or repress genes

Most bacteria using quorum sensing systems are associated with?

Plants or animals

Gram negative bacteria use what molecule for quorum sensing signals?

N-acyl homoserine lactones (AHLS)

Characteristics of N-acyl hooserine lactones (AHLs)

Small, hydrophobic, membrane -diffusible molecules

Gram positives use what type of molecule for cell signal?

Oligopeptide (5-40 aa’s)

Identify the molecules used for quorum sensing for gram positive and gram negative bacteria .

Gram negative: (AHLs) N-acyl homoserine lactones Gram positive : oligiopeptides

Quorum sensing was first discovered in?

Vibrio. Fischeri

V. Fischeri is most known for this property? It is also symbionts with marine animals most notably?

V. Fischeri is known for its bioluminescent property and notable symbionts with the bob tail squid

V. Fischeri colonizers in the light organ. Once the density is sufficient what occurs?

Bacteria turns on the lux system

Which quorum sensing system produces bioluminescence when activated?

Lux system

What are the components of the lux system?

Lux I & Lux R

What is the function of lux I

(AI synthase) is produced constitutively at low levels and is the enzyme responsible for producing the QS signal molecule, AHL(AI-1 or acyl-HSL)

In the lux system when sensitive reaches threshold concentration..

AHLs bind and activate LuxR

Once the AHL-bound LuxR dimer. List the following steps of the dimmer

1. AHL-bound Lux R dimer binds to promoter region of Lux operon 2. Binding to Lux operon positively regulates luxl gene. 3. Lux gene creates more Luxl and more AHL 4. As well down stream luxCDABEG genes

Transcription of luxCDABEG genes leads to?

The production of luciferase are and its substrate to generate bioluminescence.

What genes encode luciferase protein components in lux system?

LuxA &LuxB

Other than luxA and LuxB the lux genes encode for?

Proteins that form biosynthetic enzymes complex & long chain aldehyde complex substrate that is oxidized by luciferase

Long-chain aldehyde complex substrate that is oxidized by luciferase produces?

Blue-green light