AhmadWeek3

Part 1: Experimenting with different levels of ammonia

• Saccharomyces cerevisae, a type of yeast, uses ammonia as its primary nitrogen source
  • Past research has shown that ammonia concentration affects growth
  • In this paper, the researches wanted to know if ammonia flux, and not concentration, is the limiting factor
• Physiological Parameters
  • S. cerevisae was first grown in cultures with different levels of ammonia, but fixed glucose concentrations
  • Increases in the ammonia concentration from 29 to 61 mM showed a change in biomass from 4.9 to 8.2 g/L
  • After 61 mM, however, there was no significant increase in biomass
  • Formula to calculate ammonia flux: [dilution x (input ammonia concentration - residual ammonia concentration)/biomass]
  • At a level above 44 mM, oxygen consumption was equal to carbon dioxide production
• Ketoglutarate, Glutamate, Glutamine concentrations as ammonia concentrations were increased
  • Ketoglutarate: Decreased
  • Glutamate: Increased
  • Glutamine: Increased
• Figure 1
  • X axis: NH4 concentration
  • Y axis:
    • A: Residual NH4 concentration
    • B: 02 consumption/CO2 production
    • C: Ketoglutarate, Glutamate, Glutamine concentrations

Part 2: Gene Expression

• Northern Analyses are RNA analyses that were done to see how the changes in nitrogen affect nitrogen-regulated genes
• Figure 2 shows gene expression in response to different levels of NH4 concentrations
  • X axis: NH4 concentration
  • Y axis: Gene expression
• Permease-encoding genes:
  • GAP1
  • PUT4
• Biosynthetic genes
  • ILV5
  • HIS4
  • GDH1
  • GLN1

Part 3: Enzymes

• Figure 3 shows enzyme activity at different NH4 concentrations
  • X axis: NH4 concentration
  • Y axis: Enzyme activity
• Enzymes Studied:
  • NADPH
  • NAD-DGH
  • GS Transferase
• NADPH activity consistently decreased with increased NH4
• NAD-GDH activity consistently increased with increased NH4
• GS Transferase and Synthetase decreased slightly until 60 mM, but then leveled off with increased NH4
• The conclusion of the paper is that that the nitrogen metabolism of S. cerevisiae is regulated by ammonia concentration
  • The species may have an ammonia sensor

Word Definitions

1. Flux: "The total amount of a quantity passing through a given surface per unit time."
2. Permease:"General term for a membrane protein that increases the permeability of the plasma membrane to a particular molecule, by a process not requiring metabolic energy."
3. Oligonucleotide: "Linear sequence of up to 20 nucleotides joined by phosphodiester bonds."
4. Gram-negative:"This is a primary characteristic of bacteria that have a cell wall composed of a thin layer of peptidoglycan covered by an outer membrane of lipoprotein and lipopolysaccharide containing endotoxin."