Download Bio 226: Cell and Molecular Biology

Light-independent (dark) reactions
occur in the stroma of the chloroplast (pH 8)
Consumes ATP & NADPH from light reactions
regenerates ADP, Pi and NADP+
fixing CO2
1) RuBP binds CO2
fixing CO2
1) RuBP binds CO2
2) rapidly splits into two 3-Phosphoglycerate
• therefore called C3 photosynthesis
Reversing glycolysis
G3P has 2 possible fates
1) 1 in 6 becomes (CH2O)n
2) 5 in 6 regenerate RuBP
Reversing glycolysis
1 in 6 G3P becomes (CH2O)n
either becomes starch in chloroplast (to store in cell)
or is converted to
DHAP & exported
to cytoplasm to
make sucrose
Pi/triosePO4
antiporter only
trades DHAP for Pi
Regenerating RuBP
Basic problem: converting a 3C to a 5C compound
must assemble
intermediates that
can be broken into
5 C sugars after
adding 3C subunit
Regulating the Calvin Cycle
Rubisco is main rate-limiting step
indirectly regulated by light 2 ways
1) Rubisco activase
2) Light-induced changes in stroma
a) pH
b) [Mg2+]
c) CO2 is an allosteric activator of rubisco that only
binds at high pH and high [Mg2+]
also: stomates open in the light
Regulating the Calvin Cycle
Several Calvin cycle enzymes (e.g.Fructose-1,6bisphosphatase) are also regulated by thioredoxin
contain disulfide bonds which get oxidized in the dark
in light, ferredoxin reduces thioredoxin, thioredoxin
reduces these disulfide bonds to activate the enzyme
How light reactions talk to the Calvin cycle
SH SH
light
2Fdox
PSI
+
PSII
2e-
2Fdred
reduced
thioredoxin
S-S
oxidized
thioredoxin
S-S
oxidized
enzyme
(inactive)
SH SH
reduced
enzyme
(active)
PHOTORESPIRATION
Rubisco can use O2 as substrate instead of CO2
RuBP + O2 <=> 3-phosphoglycerate + phosphoglycolate
PHOTORESPIRATION
Rubisco can use O2 as substrate instead of CO2
RuBP + O2 <=> 3-phosphoglycerate + Phosphoglycolate
Releases CO2 without making ATP or NADH
PHOTORESPIRATION
Releases CO2 without making ATP or NADH
Called photorespiration : undoes photosynthesis
PHOTORESPIRATION
Rubisco can use O2 as substrate instead of CO2
RuBP + O2 <=> 3-phosphoglycerate + Phosphoglycolate
C3 plants can lose 25%-50% of their fixed carbon
PHOTORESPIRATION
Rubisco can use O2 as substrate instead of CO2
RuBP + O2 <=> 3-phosphoglycerate + Phosphoglycolate
C3 plants can lose 25%-50% of their fixed carbon
Both rxns occur at same active site
PHOTORESPIRATION
C3 plants can lose 25%-50% of their fixed carbon
phosphoglycolate is converted to glycolate : poison!
Detoxifying Glycolate
1) glycolate is shuttled to peroxisomes
Detoxifying Glycolate
1) glycolate is shuttled to peroxisomes
2) peroxisomes convert it to glycine
produce H2O2
Detoxifying Glycolate
1) glycolate is shuttled to peroxisomes
2) peroxisomes convert it to glycine
3) glycine is sent to mitochondria
Detoxifying Glycolate
1) glycolate is shuttled to peroxisomes
2) peroxisomes convert it to glycine
3) glycine is sent to mitochondria
4) mitochondria convert 2 glycine to 1
serine + 1 CO2
Why photorespiration loses CO2
Detoxifying Glycolate
1) glycolate is shuttled to peroxisomes
2) peroxisomes convert it to glycine
3) glycine is sent to mitochondria
4) mitochondria convert 2 glycine to 1
serine + 1 CO2
5) serine is returned to peroxisome
Detoxifying Glycolate
1) glycolate is shuttled to peroxisomes
2) peroxisomes convert it to glycine
3) glycine is sent to mitochondria
4) mitochondria convert 2 glycine to 1
serine + 1 CO2
5) serine is returned to peroxisome
6) peroxisome converts it to glycerate &
returns it to chloroplast
Detoxifying Glycolate
Why peroxisomes are next to cp and mito in C3 plants
Mitochondrion
Lipid metabolism
Most are glycerolipids: fatty acids bonded to glycerol
GLYCEROLIPIDS
Triacylglycerols = FAs on all 3 C
• store energy
GLYCEROLIPIDS
Bond FA to glycerol
Diacylglycerols = FAs on 2 Cs, headgroup on C 3
GLYCEROLIPIDS
Diacylglycerols = FAs on 2 Cs, headgroup on C 3
Form bilayers in water
Lipid metabolism
Unique aspects in
plants
Make fatty acids by
same set of reactions,
but in plastids with a
prokaryotic fatty acid
synthase
12 proteins, cf one
multifunctional
protein
Lipid metabolism
Make fatty acids in plastids with a prokaryotic FAS
• 12 proteins, instead of one multifunctional protein
• Assemble some lipids in CP, others in ER
Lipid metabolism
Make fatty acids in plastids with a prokaryotic FAS
• 12 proteins, instead of one multifunctional protein
• Assemble some lipids in CP, others in ER
• Acetyl-CoA carboxylase is also prokaryotic = 4
subunits, except in grasses (profoxydim & other grass
herbicides inhibit ACCase)
Lipid metabolism
• Acetyl-CoA carboxylase is also prokaryotic = 4
subunits, except in grasses (profoxydim & other grass
herbicides inhibit ACCase)
• Same biochem, but diff
location and enzymes
Lipid metabolism
• Acetyl-CoA carboxylase is also prokaryotic = 4
subunits, except in grasses (profoxydim & other grass
herbicides inhibit ACCase)
• Same biochem, but diff
location and enzymes
• In light cp make lots of
NADPH, and leaves are
main sinks for FA
Lipid metabolism
• Acetyl-CoA carboxylase is also prokaryotic = 4
subunits, except in grasses (profoxydim & other
herbicides inhibit ACCase)
• Same biochem, but diff
location and enzymes
• In light cp make lots of
NADPH, and leaves are
main sinks for FA
• But, each cell makes
its own FA, so NADPH
in other cells comes from
Pentose-Pi shunt
Lipid metabolism
Source of acetyl-CoA is controversial
• Most comes from plastid PDH
Lipid metabolism
Source of acetyl-CoA is controversial
• Most comes from plastid PDH
• Some comes from cytoplasmic acetate; activated in cp
• Also used to make
sterols, some amino
acids, many others
Lipid metabolism
Source of acetyl-CoA is controversial
• Most comes from plastid PDH
• Some comes from cytoplasmic acetate; activated in cp
• Also used to make sterols, some
amino acids, many others
• Why ACCase is “committed step”
Lipid metabolism
Assemble some lipids in CP, others in ER
• “16:3 plants” assemble lipids in cp using FA-ACP =
prokaryotic pathway (“primitive”)
Lipid metabolism
“16:3 plants” assemble lipids in cp using FA-ACP =
prokaryotic pathway (“primitive”)
“18:3 plants” export FA, assemble lipids in ER using
FA-CoA = eukaryotic pathway (“advanced”)
Lipid metabolism
“16:3 plants” assemble lipids in cp using FA-ACP =
prokaryotic pathway (“primitive”)
“18:3 plants” export FA, assemble lipids in ER using
FA-CoA = eukaryotic pathway (“advanced”)
Substrates for most desaturases
are lipids, not FA!
Lipid metabolism
Unique aspects in plants
Chloroplasts have lots
of galactolipids: sugar
linked directly to
diacylglycerol
Lipid metabolism
Unique aspects in plants
Chloroplasts have lots
of galactolipids: sugar
linked directly to
diacylglycerol: saves PO4
Lipid metabolism
Unique aspects in plants
Chloroplasts have lots of galactolipids: sugar linked
directly to diacylglycerol : saves PO4
A) MGDG (Monogalactosyl diacylglycerol) 50% cp
-> most abundant lipid on earth!
Lipid metabolism
Unique aspects in plants
Chloroplasts have lots of galactolipids: sugar linked
directly to diacylglycerol : saves PO4
A) MGDG (Monogalactosyl diacylglycerol) 50% cp
B) DGDG (Digalactosyl diacylglycerol) 28% cp
Lipid metabolism
Chloroplasts have lots of galactolipids: sugar linked
directly to diacylglycerol : saves PO4
A) MGDG (Monogalactosyl diacylglycerol) 50% cp
B) DGDG (Digalactosyl diacylglycerol) 28% cp
C) SQDG ( Sulphoquinovosyldiacylglycerol) 16% cp
Lipid metabolism
Chloroplasts have lots of galactolipids: sugar linked
directly to diacylglycerol : saves PO4
A) MGDG (Monogalactosyl diacylglycerol) 50% cp
B) DGDG (Digalactosyl diacylglycerol) 28% cp
C) SQDG ( Sulphoquinovosyldiacylglycerol) 16% cp
• Very unsaturated!
Lipid metabolism
Chloroplasts have lots of galactolipids: sugar linked
directly to diacylglycerol : saves PO4
A) MGDG (Monogalactosyl diacylglycerol) 50% cp
B) DGDG (Digalactosyl diacylglycerol) 28% cp
C) SQDG ( Sulphoquinovosyldiacylglycerol) 16% cp
• Very unsaturated!
• Makes membranes
very fluid
Lipid metabolism
Chloroplasts have lots of galactolipids: sugar linked
directly to diacylglycerol : saves PO4
A) MGDG (Monogalactosyl diacylglycerol) 50% cp
B) DGDG (Digalactosyl diacylglycerol) 28% cp
C) SQDG ( Sulphoquinovosyldiacylglycerol) 16% cp
• Very unsaturated!
• Makes membranes
very fluid
• Source of 3 FA
Lipid metabolism
Unique aspects in plants
1) Make fatty acids in plastids
2) large amounts of galactolipids
3) Oleosomes: oil-storing organelles with only outer leaflet
Lipid metabolism
Oleosomes: oil-storing organelles with only outer leaflet
• Put oils between the leaflets as they are made
Lipid metabolism
Oleosomes: oil-storing organelles with only outer leaflet
• Put oils between the leaflets as they are made
• Add oleosin proteins to outside: curve the membrane
Lipid metabolism
Oleosomes: oil-storing organelles with only outer leaflet
• Put oils between the leaflets as they are made
• Add oleosin proteins to outside: curve the membrane
• Oils often have unusual fatty acids
Lipid metabolism
Use fats to store carbon as well as energy!!??
• Recover C via glyoxylate cycle & gluconeogenesis
Lipid metabolism
Biological roles
• Plasma membrane lipids help
survive freezing
Lipid metabolism
Biological roles
• Plasma membrane lipids help
survive freezing
• Unacclimated cells vesiculate as
they lose water & pop when it returns
Lipid metabolism
Biological roles
• Plasma membrane lipids help
survive freezing
• Unacclimated cells vesiculate as
they lose water & pop when it returns
• Acclimated cells shrivel & reswell
Lipid metabolism
Biological roles
• Plasma membrane lipids help survive freezing
• Mito lipid composition may also influence chilling
sensitivity
• CS plants (eg bananas) are damaged at 10˚ C
Lipid metabolism
log respiration (nmol O2/min/mg protein)
Biological roles
• Plasma membrane lipids help survive freezing
• Mito lipid composition may also influence chilling
sensitivity
• CS plants (eg bananas) are damaged at 10˚ C
• Mito show defects at <10˚ C not seen in other plants
3.0
B) SB-H mitochondria
A) fr2 x fr1 mitochondria
34°
2.8
2.6
18°
2.4
35°
25°
24°
2.2
2.0
16°
Control
17:0
18:1
Control
17:0
18:1
1.8
3.15 3.20 3.25 3.30 3.35 3.40 3.45 3.50 3.55 3.15
3.60 3.20 3.25 3.30 3.35 3.40 3.45 3.50 3.55 3.60
1/T (°K x 1000)
1/T (°K x 1000)
Lipid metabolism
CS plants (eg bananas) are damaged at 10˚ C
• Mito show defects at <10˚ C not seen in other plants
• Membrane lipids show phase changes at these T
Lipid metabolism
CS plants (eg bananas) are damaged at 10˚ C
• Mito show defects at <10˚ C not seen in other plants
• Membrane lipids show phase changes at these T
• Blamed on saturated PG
log respiration (nmol O2/min/mg protein)
Lipid metabolism
Biological (& commercial) roles
• Plasma membrane lipids help survive freezing
• Mito lipid composition influences chilling sensitivity
• Mito show defects at <10˚ C not seen in other plants
• unsaturated FA did not fix CS, but saturated FA
made it worse: reason for GM desaturases
3.0
B) SB-H mitochondria
A) fr2 x fr1 mitochondria
34°
2.8
2.6
18°
2.4
35°
25°
24°
2.2
2.0
16°
Control
17:0
18:1
Control
17:0
18:1
1.8
3.15 3.20 3.25 3.30 3.35 3.40 3.45 3.50 3.55 3.15
3.60 3.20 3.25 3.30 3.35 3.40 3.45 3.50 3.55 3.60
1/T (°K x 1000)
1/T (°K x 1000)
Lipid metabolism
Other commercial aspects
• Yield and quality of seed oil is very important:12
million tons/year
Lipid metabolism
Other commercial aspects
• Yield and quality (especially unsaturation) of seed oil is
very important:12 million tons/year
• Want more double bonds, especially -3, for health
• Want less double bonds for shelf life and taste
• Each double bond increases p(oxidation) 40x
Lipid metabolism
Other commercial aspects
• Yield and quality (especially unsaturation) of seed oil is
very important:12 million tons/year
• Want more double bonds, especially -3, for health
• Want less double bonds for shelf life and taste
• Each double bond increases p(oxidation) 40x
• Have GM oils with more & less double bonds
Lipid metabolism
Other commercial aspects
• Yield and quality of seed oil is very important:12
million tons/year
• Also have markets for many specialized oils
Lipid metabolism
Other commercial aspects
• Yield and quality of seed oil is very important
• Also have markets for many specialized oils
• Have genetically-engineered many crops to alter
seed oils or produce specific fats
Lipid metabolism
Biofuels are now very fashionable
• Biodiesel = fatty acid methyl esters
• Trans-esterify oils to make them volatile
Lipid metabolism
Biofuels are now very fashionable
• Biodiesel = fatty acid methyl esters
• Trans-esterify oils to make them volatile
• Projected to be 10% of european diesel in 2015
Lipid metabolism
Biofuels are now very fashionable
• Biodiesel = fatty acid methyl esters
• Trans-esterify oils, used cooking oil, etc
• Projected to be 10% of european diesel in 2010
• Also use coconut & other oils directly in diesel engines
Lipid metabolism
Also use coconut & other oils directly in diesel engines
• Just need to be sufficiently fluid to reach cylinder
Lipid metabolism
Also use coconut & other oils directly in diesel engines
• Just need to be sufficiently fluid to reach cylinder
• Add double bonds to fatty acids or make them shorter