Cycle 5: Integrated Metabolism

• Measuring Respiration(looks at the rate of oxygen consumption)

  Use O2 electron chamber (break chlamy cell, purify intact mitochondria, wash mitochondria, incubate isolated mitochondria with buffer)
• Speed of O2 consumption

  = Speed of Respiration
• Mitochondria involved with electron transport chain

  Starts consumption if both it and NADH present
• NADH involved with electron transport chain

  Starts consumption if both it and mitochondria are present
• ADP + Pi involved with electron transport chain

  Speeds up O2 consumption, only if NADH and mitochondria are present
• Uncoupler involved with electron transport chain

  Speeds up O2 consumption only if NADH and mitochondria are present
• The steeper the slope (O2 consumption curve)

  The faster the rate of oxygen consumption
• ADP and Pi (respiratory control)

  Proton entry through ATP synthase. Easier for ETC to pump H+. Higher O2 consumption.
• Uncoupler

  Dissipates the proton gradient, ETC speeds up. Highest rate of O2 consumption
• G3P from the calvin cycle is sourced for

  Energy through respiration. Biosynthetic reactions
• Cell Growth


  One cell divides into two cells. Use G3P as a source of carbon for the backbones of molecules needed for cell growth (ex: nucleic acids, amino acids, fatty acids, etc)
• Autotrophic + heterotrophic


  = Mixotrophic
• Heterotrophic


  Growth is where we take in external food sources and there is no photosynthesis. This relies on acetate and when there is no light, acetate transporters are present to be used
• Autotrophic


  Growth through photosynthesis and glucose synthesis. When there is no light present, we cannot undergo photosynthesis and there are no glucose transporters to use glucose
• Mixotrophic


  Growth is when there is the presence of light and acetate in the environment. Chlamy has the highest growth rate because it can get growth through photosynthesis and using acetate
• Photosynthesis involves gas exchange


  CO2 is consumed, O2 is produced
• How to measure photosynthesis


  Put a culture of chlamy into CO2 analyzer, expose the culture to varying light intensities, access changes in CO2 levels
• Rate of Respiration (light response curve)


  No light = no photosynthesis, but cellular respiration is nt light-dependent, net CO2 release (constant rate)
• Light compensation curve (light response curve)


  Rate of carbon loss (respiration) is equal to rate of carbon gain (photosynthesis), therefore the y-axis indicated net rate of carbon fixation
• Light -limited region (light response curve)


  There is an approximate linear relationship between light intensity and rate of photosynthesis. When there is net carbon gain = growth
• Light saturated point (light response curve)

  Rate of photosynthesis reaches its max. Pmax = maximum net rate of CO2 fixation (does not equal maximum rate of CO2 fixation). Limited by maximum turnover rate of enzymes in the calvin cycle, not substrates
• Net photosynthesis = Pmax

  = Gross photosynthesis - respiration (this is a measure of carbon gain)
• What limits the rate of carbon fixation as a function of light?


  Enzymes: rubisco has a limit to how fast it can turnover product, even with an abundance of NADPH and ATP. Available CO2
• No substrate (enzyme kinetics)


  = zero velocity
• Increasing velocity (enzyme kinetics)


  Is due to increasing [S]
• Km (enzyme kinetics)


  Is a measure of affinity. It is the [S] at which half of maximum velocity is reached
• Vmax (enzyme kinetics)


  Is a measure of the max velocity
• Low Km =


  High affinity for substrate
• High Km = 


  Low affinity for substrate
• Order of affinity


  Km1 > Km2 > Km3
• Inhibitor(competitive inhibition)


  Resembles original substrate, binds to active site, binding can be reversible or irreversible
• Effect (competitive inhibition)


  = increased Km (no change in 1/2 Vmax or Vmax)
• Increasing [S] can ...


  Reduce effects of reversibly-bound inhibitors
• Advantage of enzyme regulation in non-competitive (allosteric) regulation


  It is faster to regulate at the level of enzyme activity vs transcription or translation
• Inhibitor (non-competitive inhibition curve)


  Does not bind to the active site (binds elsewhere), causes a conformational change that makes the substrate unable to bind
• Effect (non-competitive inhibition curve)


  Change in 1/2 Vmax or Vmax (no change in Km)