25th International Meeting on Organic Geochemistry IMOG 2011

P-420
� 2 H differences among lipids synthesised via the ACT
(acetogenic), MVA (mevalonic) and DXP (1-deoxy-D-xylulose 5phosphate)
pathways in higher plant leaves: possible evidence
for intracellular water (H+) isotopic heterogeneity
Youping Zhou 1,2 , Kliti Grice 1 , Hilary Stuart-Williams 2 , Graham Farquhar 2 , Charles Hocart 2
1 Curtin University, Perth, Australia, 2 Australian National University, Canberra, Australia (corresponding
author:y.zhou@curtin.edu.au)
Carbon-bound hydrogen atoms in lipids from higher
plant leaves have three immediate sources: 1)
inherited from biosynthetic precursors, 2) water (H + ) in
which biosynthesis occurs and 3) bio-reductant
NAD(P)H [1,2,3,4,5] . Hydrogen isotopic ratios ( 2 H/ 1 H) of
lipids are determined by those of the sources modified
by isotopic fractionations during biosynthesis. Lipids
synthesised via the ACT pathway (e.g. n-alkanes), the
MVA (e.g. sterols) and the DXP pathway (e.g. phytol)
were isolated from leaves of C3 and C4 species
grown under controlled and field conditions.
Compound-specific isotopic analysis (CSIA) of these
lipid revealed that an inter-pathway hydrogen isotopic
enrichment order: � 2 HACT > � 2 HMVA > � 2 HDXP exists in
all species examined [1,3,5] . There is an isotopic
enrichment ordering between photosynthetic modes:
� 2 HC4 < � 2 HC3 (for ACT), � 2 HC4 > � 2 HC3 (for MVA and
DXP) [1,3,5] . Average inter-pathway differences for C4
plants are smaller than for C3 plants.
An investigation of the histories of the individual
hydrogen atoms in these lipid molecules, based on
available knowledge of their biosynthetic chemistry,
indicated that the observed inter-pathway order is a
result of intracellular NADPH variation and
intracellular water isotopic heterogeneity. NADPH and
water (H + ) in the chloroplast are isotopically more
depleted in 2 H than their respective cytosolic
counterparts. The inter-compartmental water and
NADPH isotopic differences are passed onto lipid
molecules synthesised in each cellular compartment
via enzyme-catalysed hydration or NAD(P)H
reduction of metabolic intermediates.
Compartmentalisation of initial photosynthetic CO2
fixation into the C4 mesophyll (M) cell and final CO2
fixation in the bundle sheath (BS) cell results in more
2 H-depleted pyruvate (in comparison to C3), the
precursor for lipids synthesised via the ACT pathway
in the chloroplast, due to the exchange of carbonbound
hydrogen in pyruvate with the less enriched
cellular water in the BS cell (relative to M cellular
water) during the malate-pyruvate shuttle.
Compartmentalisation in C4 plants also results in less
depleted NADPH in the M chloroplast (in comparison
to C3) for lipid synthesis via DXP pathway due to the
selective export of 2 H-depleted NADPH from the M
cell to the BS cell and a less depleted pyruvate in the
BS cytosol for lipid synthesis via MVA pathway due to
the suppression of photorespiration (Fig 1) [5] .
Mesophyll chloroplast
malate
malate
NADP +
NADPH
pyruvate
Bundle sheath cytosol
COO
O
-
H �
H �
H b
COO
OH
-
H �
H �
O - 2e + H OPi RuBP
GAP O
H C5 H
H C3C HO HO O
C4 H
C3 H
C2 H
H
C24 OH
C H
15
OPi H
OC
H C1 H
OPi HO
CO2 H
H
FBP
O
OH
C3C H C3C H G6P
C24 OH
C24 O
Starch
C15 H
H C15 H
OPi DHAP Maltose
OPi + + NADP + NADPH + H +
H
+ -
H2O H + HO
Calvin Cycle
+
+ -
H2O H + HO
H
lumen
sterol
O
HO
H
HO
HO
C15 H
C24 H
C3C OH
C3C H
C24 H
+
H + H +
H +
H +
H +
H +
H +
H + H +
H +
H +
H +
H +
H +
stroma
PS I
Cyt bf
Cyt bf
pyruvate
malate pyruvate
PS I
PGA
PGA
NADPH NADP +
DHAP Maltose
DHAP
COO
OPi -
H �
H �
O
COO
CO2 -
H �
H �
O - COO
O
OC
-
H �
H �
O - NADP
OC
+
COO
OH
NADPH
-
H �
H �
O - H
OC
COO
OPi
-
H ��b
H ��b
COO
O
ATP
AMP
-
H �
H �
H b
malate pyruvate
PGA OH H
O C3C C24 C15 OPi OH H H
PS I
PS II
H
Cyt bf
OAA
PEP
GAP OH H
OPi O C3C C24 C
H
15
H H H
TPI H
OH Cyt bf
C3C H
C24 O
PS II
C15 H
PS I
C18 Fatty acid
Mesophyll cytosol
+
H +
H +
H +
H +
H +
H +
H +
H +
H +
H +
H +
PGA
DHAP
Fig 1. Compartmentalisation of C4 photosynthesis into
mesophyll cell and bundle sheath cell.
[1] Chikaraishi et al. (2009) Phytochem 70, 569-573
[2] Schmidt et al. (2008) Phytochem Rev 2, 61-85
[3] Sessions et al. (1999) Org Geochem 30, 1193-1200
[4] Zhang et al. (2009) PNAS 106, 12580-12586
[5] Zhou et al. (2010) Phytochem 71, 388-403
stroma
lumen
2H 2 O 4H + + O 2 + 4e
Bundle sheath chloroplast
H 2 O H + + HO -
H
glycolysis TCA
C 15 H
OP i
H 2 O H + + HO -
2e + H + + NADP + NADPH + H +
glucose
548