Bacterial Metabolism - Richardson Flashcards
How do cells make energy available from nutrients?
Electron carrying groups. There are 2e- carriers like NADH and FAD. NAD+ accepts electrons becoming NADH and same for FAD to FADH2. They both carry high energy electrons for energy transport. There are also 1e- carriers such as heme where iron is an essential e- donor. Bacteria can utilize many mechanisms for e- transport.
Explain Chemiosmotic Theory and proton motive force in bacteria
ATP synthesis (energy production) stems from the electrochemical gradients present between inner membranes and periplasmic space in bacteria. Using energy from e- carrying groups, protons are pumped out of the inner membrane to the periplasm to establish a charge gradient. This charge gradient is also referred to as PMF. There is also a pH gradient caused by efflux of H+.
Ex. PMF provides free energy utilized by flagella to do mechanical work.
Describe bacterial fermentation
In the absence of oxygen and e- transport, some bacteria can ferment lactose utilizing electrons to reduce pyruvate to lactate.
Explain biochemical logic
Easier to break double bonds next to oxygen atoms due to resonance. Explains preference for glucose during respiration.
Steps Involved in Metabolomics
- Quenching
- Extract
- Identify
- Quantify
- Flux Determination
Explain quenching procedure in metabolomics
Procedure to arrest metabolism in cells before turnover of metabolites under investigation. Utilizes mixtures of organic solvent such as acetonitrile and methanol.
Explain the extraction of metabolites in metabolomics
- Cell disruption by mechanical, enzymatic or chemical means.
- Removing cell debris by centrifugation
- Concentrating
Explain the identification of metabolites in metabolomics
LC/MS and GC/MS
Explain how to quantify metabolites in metabolomics
NMR –> Mass Spec is not quantitative
Flux Determination
- Linear algebra
- Labeled metabolites measured by MS and NMR.
Ex. You can use C13 to tag glucose to pyruvate. If you go through glycolysis you get singly labeled carbons, if you go through fermentation you get doubly labeled carbons.
Briefly outline basic bacterial metabolic needs
Bacterial cells need to be able to utilize energy locked in small molecules such as glucose. This is done by oxidation of glucose and transfer of high energy e- via e- transport chains to generate ATP.
Briefly outline redox and group transfer potentials
Redox reactions are necessary metabolic processes that unlock energy stored in molecules and subsequent group transfer of e- for energy. For example oxidation of glucose results in pyruvate which its derivatives donate e- to NAD+ and FAD+.
Briefly outline chemiosmotic theory
Proton gradients create a charge gradient sufficient to do cellular work.
Ex. ATP synthesis and flagella movement
Briefly outline nutrient acquisition in bacteria
Bacteria can utilize several metabolic pathways to catabolize carbon sources and metabolic precursors.
Briefly outline respiration vs fermentation
Respiration is e- conservative = more energy efficient. Fermentation dissipates free e- and is less energy efficient.
Describe batch culture in bacteria
Growing bacteria in a flask. Undergo a lag phase, followed by a exponential growth phase before reaching a stationary phase. Stationary phase is based on physical and nutrient constraints in the environment. Eventually, after resources are used up, death and decline will occur.
Describe continuous growth culture in bacteria
Maintaining maximum growth rate for extended periods of time. Achieved by consistently diluting bacterial cultures and refreshing nutrients in the media.
Describe carbon catabolite regulation in gram negative bacteria
Phosphorylated E2A signals that there is no available glucose for consumption. E2A turns on adenylate cyclase which activates CRP txn factor to induce gene expression for alternative sugar consumption in a sequential manner.
Describe carbon catabolite regulation in gram positive bacteria
Fructose bisphosphate accumulation signals no glucose. Activates txn factors for any alternative sugar. Much faster response to environment.
What is gram positive anti-termination?
Stem loops that form to prevent transcription
How do bacteria store Fe?
They lock Fe away in ferratin so it is innert
Describe host-iron sequestration
Multiple layers of binding up Fe so that it cannot be utilized by bacteria. Pathogens have evolved to scavenge iron.
What regulates iron uptake in bacteria?
Ferric uptake regulator (FUR).
What does a low Km mean?
High substrate affinity
What does a high Km mean?
Low substrate affinity
What is Vmax?
The maximum rate of an enzymatic reaction.
What happens to Vmax and Km during competitive inhibition?
Vmax remains the same but Km changes –> Active site is bound so enzyme affinity is locked. With enough substrate, rxn can proceed at same rate.
What happens to Vmax and Km during non-competitive inhibition?
Enzyme has the same affinity for its substrate, but is slower. Therefore, Vmax changes but not Km.
