viridis Reaction Center Architecture Type II photosystems of higher plants, green algae, and cyanobacteria contain more than 20 subunits and are considerably more complex than the R.. Ea
Trang 1phyll (BChl) The eis then transferred via the L bacteriopheophytin (BPheo) to
QA, which is also an L prosthetic group The corresponding site on M is occupied by
a loosely bound quinone, QB, and electron transfer from QAto QBtakes place An
interesting aspect of the system is that no electron transfer occurs through M, even
though it has components apparently symmetric to and identical to the L etransfer
pathway
The reduced quinone formed at the QBsite is free to diffuse to a neighboring
cytochrome bc1membrane complex, where its oxidation is coupled to H
transloca-tion and, hence, ultimately to ATP synthesis The use of light energy to drive ATP
synthesis by the concerted action of these membrane proteins is called
photophos-phorylation (Figure 21.15)
Cytochrome c2, a periplasmic protein, serves to cycle electrons back to P870via
the four hemes of the reaction center cytochrome subunit A specific tyrosine
residue of L (Tyr162) is situated between P870 and the closest cytochrome heme
This Tyr is the immediate edonor to P870and completes the light-driven
elec-tron transfer cycle
The Molecular Architecture of PSII Resembles the R viridis Reaction
Center Architecture
Type II photosystems of higher plants, green algae, and cyanobacteria contain
more than 20 subunits and are considerably more complex than the R viridis
reaction center The structure of PSII from the thermophilic cyanobacterium
Light Outside
Cytoplasm
Cyt c2
Cyt c2
QH2
QB QA
L M
H
2 H +
Fe
2 H +
4 H +
4 H +
4 H +
Bacterial
F1F0–ATP
synthase
+ Pi
2e–
Q
2 H +
Cyt b/c1
ATP ADP
ACTIVE FIGURE 21.15 Photophosphorylation Photoexcitation of the R viridis RC leads to
reduction of a quinone, Q, to form QH 2 Oxidation of QH 2by the cytochrome bc1 complex leads to H
trans-location for ATP synthesis by the R viridis F1 F 0–ATP synthase Test yourself on the concepts in this figure at
www.cengage.com/login.
Trang 2644 Chapter 21 Photosynthesis
Synechococcus elongatus has been revealed by X-ray crystallography, providing insight
into PSII structures in general Interestingly, both type II and type I photosystems
show significant similarity to the R viridis reaction center, thus establishing a
strong evolutionary connection between reaction centers
S elongatus PSII is a homodimeric structure Each “monomer” has a mass of
almost 350 kD and 23 different protein subunits, the 4 largest being the reaction
center pair of subunits (D1 and D2) and two chlorophyll-containing inner antenna subunits (CP43 and CP47) that bracket D1 and D2 (Figure 21.16) Together, CP43
and CP47 have a total of 26 Chl a molecules, and exciton energy is collected and
transferred from them to P680 Collectively, the protein subunits in a PSII
“monomer” have at least 34 transmembrane -helical segments, 22 of which are
found in the D1-D2-CP43-CP47 “core” structure D1 and D2 each have five membrane-spanning-helices Structurally and functionally, these two subunits are
a direct counterpart of the L and M subunits of the R viridis reaction center P680 consists of a pair of Chl a molecules, with D1 and D2 each contributing one D1 and
D2 each have two other Chl a molecules, one near each P680 (ChlD1 and Chl D2 , re-spectively) and another that interacts with CP43/CP47 (ChlZ-D1and ChlZ-D2, respec-tively) (Figure 21.16) Two equivalents of pheophytin (Pheo) are located on D1 and D2 The tyrosine species D is Tyr161in the D1 amino acid sequence Complexed to D2 is a tightly bound plastoquinone molecule, QA Electrons flow from P680* to ChlD1and on to PheoD1 PheoD1then transfers the electron to QAon D2, where it then moves to a second plastoquinone situated in the QBsite on D1 (Figure 21.16) Electron transfer from QA and QB is assisted by the iron atom located between them Each plastoquinone that enters the QB site accepts two electrons derived from water and two Hfrom the stroma before it is released into the membrane as the hydroquinone PQH2 Thus, two photons are required to reduce each PQ that enters the QB site The stoichiometry of the overall reaction catalyzed by PSII is
2 H2O 2 PQ 4 h ⎯⎯→ O2 2 PQH2 The (Mn)4complex is located on the
Fe
PD1 PD2
Z-D2
(Mn)4
Stroma
Thylakoid lumen
(a)
(b)
FIGURE 21.16 Molecular architecture of the
Synechococ-cus elongatus PSII dimer (a) The arrow shows the path of
electron transfer from P680* to Chl D1 to Pheo D1 to Q A
on D2 and then, via the Fe atom, to Q B on D1 The Tyr 161
residue of D1, symbolized by Y z , is situated between
P680 and the (Mn) 4cluster (b) Structure of S elongatus
PSII (pdb id 1S5L) Chlorophylls of the reaction center
and electron transfer path are shown in green;
pheo-phytins, in blue The OECs are shown in brick red.
(Adapted from Barber, J., 2003 Photosystem II: The engine of life.
Go to CengageNOW and click
CengageInteractive to explore the R viridis reaction
center, a complex scaffold for transduction of light
energy.
Trang 3lumenal side of the thylakoid membrane Thus, protons liberated from H2O
mole-cules at the Mn site are deposited directly into the lumen
How Does PSII Generate O2from H2O?
PSII catalyzes what is probably the most thermodynamically difficult reaction in
na-ture, the light-driven oxidation of water to molecular oxygen The protons and
elec-trons released in this reaction are used to reduce NADPto NADPH and to
estab-lish a proton gradient across the photosynthetic membrane that can be tapped to
drive chemiosmotic ATP synthesis (see Figure 21.20) Accumulation of molecular
oxygen in the atmosphere as a by-product of the photo-oxidation of water has
trans-formed the planet since the evolutionary appearance of this reaction some 2 billion
years ago in cyanobacteria How does PSII evolve oxygen?
The Structure of the Oxygen-Evolving Complex The architecture of the S
elon-gatus photosynthetic OEC reveals a large globular protein domain juxtaposed on the
lumenal side of the D1 subunit of PSII (Figure 21.16) The active site of the OEC
contains a cubelike metal cluster that consists of four manganese ions, one calcium
ion, and five oxygen atoms bridging the Mn atoms, as shown in Figure 21.17 This
metal cluster is held by Glu189, Asp342, His332, and His337of the PSII D1 subunit and
Glu354of CP43 Chloride ion (Cl) is required for O2evolution, and Clis believed
to be a Ca2ligand Note also that Tyr161(Y Z) is situated near the metal cluster,
ide-ally poised to serve in electron transfer between H2O and P680 When four ehave
been removed from the cluster (one from each Mn atom) through etransfer to
PSII via Tyr161, two H2O molecules provide the four eneeded to re-reduce the Mn
atoms and O2is evolved The four Hreleased contribute to the proton gradient
The Molecular Architecture of PSI Resembles the R viridis Reaction
Center and PSII Architecture
The structure of PSI from the cyanobacterium S elongatus also has been solved
by X-ray crystallography, completing our view of reaction center structure and
confirming the fundamental similarities in organization that exist in these
energy-transducing integral membrane proteins Because of direct correlations with
infor-mation about eukaryotic PSI, this cyanobacterial PSI provides a general model for all
P700-dependent photosystems
S elongatus PSI exists as a cloverleaf-shaped trimeric structure Each “monomer”
(356 kD) consists of 12 different protein subunits and 127 cofactors: 96 chlorophyll a
molecules, 2 phylloquinones, 3 Fe4S4clusters, 22 carotenoids, and 4 lipids that are an
intrinsic part of the protein complex (Figure 21.18) All of the electron-transferring
prosthetic groups essential to PSI function are localized to just three polypeptides:
PsaA, PsaB, and PsaC PsaA and PsaB (83 kD each) compose the reaction center
het-erodimer, a structural pattern now seen as universal in photosynthetic reaction centers
PsaA and PsaB each have 11 transmembrane -helices, with the 5 most C-terminal
-helices of each serving as the scaffold for the reaction center photosynthetic
electron-transfer apparatus PsaC interacts with the stromal face of the PsaA–PsaB heterodimer
PsaC carries the two Fe4S4clusters, FAand FB, and interacts with PsaD Together they
provide a docking site for ferredoxin The electron-transfer system of PSI consists of
three pairs of chlorophyll molecules: P700 (a heterodimer of Chl a and an epimeric
form, Chl a) and two additional Chl a pairs (symbolized by A0) that mediate e
trans-fer to the quinone acceptor The S elongatus quinone acceptor (A1 ) is phylloquinone
(also known as vitamin K1) The Fx Fe4S4cluster bridges PsaA and PsaB; two of its four
cysteine ligands come from PsaA, the other two from PsaB Photochemistry begins
with exciton absorption at P700, almost instantaneous electron transfer and charge
separation (P700⬊A0 ), followed by transfer of the electron from A0to A1and on
to FXand then FAand FB, where it goes on to reduce a ferredoxin molecule at the
“stromal” side of the membrane The positive charge at P700and the eat FA/FB
represent a charge separation across the membrane, an energized condition created
His332 Asp 170
Asp342 CP43
Glu354 Glu 333
Tyr 161
His 190
His 337
Gln165 Glu189
MnA
MnB
MnC
MnD Ca
Ala 344 (C-term)
FIGURE 21.17Structure of the PSII oxygen-evolving complex (OEC) Four Mn atoms (red, lettered A–D) and a
Ca atom (green) form the water-splitting metal cluster
of the OEC Bridging O atoms are purple (Adapted from Figure 4 in Yano, J., et al., 2006 Where water is oxidized to dioxy-gen: Structure of the photosynthetic Mn 4 Ca cluster.
Science 314:821–825.)
Trang 4646 Chapter 21 Photosynthesis
FeS A FeS B
FeS X
P700
A1 O
O
O O
A0
Plastocyanin
Plastocyanin docking
Lumen
PsaC PsaD
Ferredoxin docking
Ferredoxin
PsaF
FIGURE 21.18 The molecular architecture of PSI (a) Subunit organization.(Adapted
from Golbeck, J H., 1992 Structure and function of photosystem I Annual Review of Plant
Physi-ology and Plant Molecular BiPhysi-ology 43:293–324; and Fromme, P., Jordan, P., and Krausse, N., 2001.
Structure of photosystem I Biochimica Biophysica Acta 1507:5–31.)(b) Molecular graphic of
theoretical model for PSI PsaA is orange; PgaB is magenta; PsaC is yellow The iron-sulfur clusters are red (pdb id 1YO9).
FIGURE 21.19 View of the plant PSI-LHC1
supercom-plex, from the stromal side of the thylakoid membrane.
ChI molecules are shown in green and carotenoids
and lipids in red The 16 protein subunits are shown as
ribbon diagrams in the background, with the positions
of PsaG, PsaH, PsaK, and LHC1-4 subunits indicated.
(Adapted from Figure 4 in Nelson, N., and Yocum, C F., Structure
and function of photosystems I and II Annual Review of Plant
Trang 5by light Plastocyanin (or in cyanobacteria, a lumenal cytochrome designated
cyto-chrome c 6) delivers an electron to fill the electron hole in P700
How Do Green Plants Carry Out Photosynthesis?
Do the higher plant photosystems follow the structural and functional pattern
first revealed in the bacterial RC and recapitulated in the cyanobacterial PSI and
PSII? With new structural information for the higher plant PSI (from Pisum
sativum, garden pea) and PSII (from spinach), the fundamental organization
pattern for photosystems seen earlier is confirmed for higher plants Further, the
structure of a plant membrane protein supercomplex consisting of the PSI
reac-tion center and its light-harvesting antenna LHC1 (light-harvesting complex 1)
has been described This supercomplex exists as a “monomer” composed of 16
distinct protein subunits and about 200 prosthetic groups, including 167
chloro-phylls, 2 phylloquinones, and 3 Fe4S4clusters (Figure 21.19) The four LHC1
subunits form an arc around one side of the PSI RC A second light-harvesting
complex (LHC2) binds to another side This plant PSI system, like all
photosys-tems, is remarkably efficient, showing a quantum efficiency of nearly 1 (one
elec-tron transferred per photon falling anywhere within the supercomplex) The
many Chl and other light-harvesting molecules of the supercomplex form an
in-tegrated network for highly efficient transfer of light energy into P700
The quantum yield of photosynthesis can be defined as the amount of product
formed per equivalent of light input In terms of exciton delivery to reaction center
Chl dimers and subsequent etransfer, the quantum yield of photosynthesis typically
approaches the theoretical limit of 1 The quantum yield of photosynthesis can also
be expressed as the ratio of CO2fixed or O2evolved per photon absorbed
Interest-ingly, an overall stoichiometry of three Htranslocated into the thylakoid vesicle has
been observed for each electron passing from H2O to NADP Two photons per
cen-ter would allow a pair of electrons to flow from H2O to NADP(see Figure 21.11),
resulting in the formation of 1 NADPH and 1
2 O2 More appropriately, 4 h per
cen-ter (8 quanta total) would drive the evolution of 1 O2, the reduction of 2 NADP,
and the translocation of 12 H Current estimates suggest that 3 ATPs are formed for
every 14 Htranslocated, so (12/14)3 2.57 ATP would be synthesized from an
in-put of 8 quanta
The energy of a photon depends on its wavelength, according to the equation
E h hc/, where E is energy, c is the speed of light, and is its wavelength
Ex-pressed in molar terms, an Einstein is the amount of energy in Avogadro’s number
of photons: E Nhc/ Light of 700-nm wavelength is the longest-wavelength and
the lowest-energy light acting in the eukaryotic photosystems An Einstein of
700-nm light is equivalent in energy to approximately 170 kJ Eight Einsteins of
this light, 1360 kJ, theoretically generate 2 moles of NADPH, 2.57 moles of ATP, and
1 mole of O2
Calculation of the Photosynthetic Energy Requirements for Hexose
Synthesis Depends on Hⴙ/h and ATP/HⴙRatios
The fixation of carbon dioxide to form hexose, the dark reactions of photosynthesis,
requires considerable energy The overall stoichiometry of this process (see Equation
21.3) involves 12 NADPH and 18 ATPs Thus, the ATP/NADPH ratio for CO2fixation
is 1.5 To generate 12 equivalents of NADPH necessitates the consumption of 48
Ein-steins of light, minimally 170 kJ each However, if the preceding ratio of 1.29 ATPs per
NADPH is correct, only 15.5 or so ATPs would be produced for CO2fixation To make
up the deficit of 2.5 ATPs would require 35 Hor about 12 more etransferred from
HO to NADP(an additional 24 Einsteins) From 72 Einsteins, or 12,240 kJ, 1 mole
Trang 6648 Chapter 21 Photosynthesis
of hexose would be synthesized The standard free energy change, G°, for hexose
formation from carbon dioxide and water (the exact reverse of cellular respiration)
is2870 kJ/mol Note that many assumptions underlie these calculations, including assumptions about the ATP/Hratio, the H/eratio, and ultimately, the relation-ship between quantum input and overall yields of NADPH and ATP Also, cyclic pho-tophosphorylation (see later section titled Cyclic Phopho-tophosphorylation Generates ATP but Not NADPH or O2) leads to ATP synthesis and may aid in making up the ATP deficit just mentioned
Light-driven ATP synthesis, or photophosphorylation, is a fundamental part of the photosynthetic process The conversion of light energy to chemical energy results
in electron-transfer reactions, which lead to the generation of reducing power (re-duced quinones or NADPH) Coupled with these electron transfers, protons are driven across the thylakoid membranes from the stromal side to the lumenal side These proton translocations occur in a manner analogous to the proton transloca-tions accompanying mitochondrial electron transport that provide the driving force for oxidative phosphorylation (see Chapter 20) Figure 21.11 indicates that proton translocations can occur at a number of sites For example, protons are produced
in the thylakoid lumen upon photolysis of water by PSII The oxidation–reduction events as electrons pass through the plastoquinone pool and the Q cycle are an-other source of proton translocations The proton transfer accompanying NADP reduction also can be envisioned as protons being taken from the stromal side of the thylakoid vesicle The current view is that three protons are translocated for each electron that flows from H2O to NADP Because this electron transfer re-quires two photons, one falling at PSII and one at PSI, the overall yield is 1.5 pro-tons per quantum of light
The Mechanism of Photophosphorylation Is Chemiosmotic
The thylakoid membrane is asymmetrically organized, or “sided,” like the mito-chondrial membrane It also shares the property of being a barrier to the passive diffusion of Hions Photosynthetic electron transport thus establishes an electro-chemical gradient, or proton-motive force, across the thylakoid membrane with the interior, or lumen, side accumulating Hions relative to the stroma of the chloro-plast Like oxidative phosphorylation, the mechanism of photophosphorylation is chemiosmotic
A proton-motive force of approximately 250 mV is needed to achieve ATP syn-thesis This proton-motive force, p, is composed of a membrane potential, , and
a pH gradient, pH (see Chapter 20) The proton-motive force is defined as the
free energy difference, G, divided by Ᏺ, Faraday’s constant:
p G/Ᏺ (2.3 RT/Ᏺ)pH (21.5)
In chloroplasts, the value of is typically 50 to 100 mV and the pH gradient is
equivalent to about 3 pH units, so (2.3 RT/Ᏺ)pH 170 mV This situation
con-trasts with the mitochondrial proton-motive force, where the membrane potential contributes relatively more to p than does the pH gradient.
CF1CF0–ATP Synthase Is the Chloroplast Equivalent
of the Mitochondrial F1F0–ATP Synthase
The transduction of the electrochemical gradient into the chemical energy repre-sented by ATP is carried out by the chloroplast ATP synthase, which is highly analo-gous to the mitochondrial F1F0–ATP synthase The chloroplast enzyme complex is
called CF 1 CF 0 –ATP synthase, “C” symbolizing chloroplast Like the mitochondrial complex, CFCF–ATP synthase is a heteromultimer of
Trang 7c-subunits (see Chapter 20), consisting of a knoblike structure some 9 nm in
diame-ter (CF1) attached to a stalked base (CF0) embedded in the thylakoid membrane The
mechanism of action of CF1CF0–ATP synthase in coupling ATP synthesis to the
col-lapse of the pH gradient is similar to that of the mitochondrial ATP synthase
de-scribed in Chapter 20 However, higher plant CF1CF0–ATP synthase is believed to have
14 c-subunits in its F0rotor, implying that one turn of F0would require 14 Hand lead
to synthesis of 3 ATPs The mechanism of photophosphorylation is summarized
schematically in Figure 21.20
Photophosphorylation Can Occur in Either a Noncyclic or a Cyclic Mode
Photosynthetic electron transport, which pumps Hinto the thylakoid lumen, can
occur in two modes, both of which lead to the establishment of a transmembrane
proton-motive force Thus, both modes are coupled to ATP synthesis and are
con-sidered alternative mechanisms of photophosphorylation, even though they are
dis-tinguished by differences in their electron transfer pathways The two modes are
cyclic and noncyclic photophosphorylation Noncyclic photophosphorylation has
been the focus of our discussion and is represented by the scheme in Figure 21.20,
where electrons activated by quanta at PSII and PSI flow from H2O to NADP, with
concomitant establishment of the proton-motive force driving ATP synthesis Note
that in noncyclic photophosphorylation, O2is evolved and NADPis reduced
Cyclic Photophosphorylation Generates ATP but Not NADPH or O2
In cyclic photophosphorylation, the “electron hole” in P700created by electron
loss from P700 is filled not by an electron derived from H2O via PSII but by a cyclic
pathway in which the photoexcited electron returns ultimately to P700 This
path-way is schematically represented in Figure 21.11 by the dashed line connecting
ferredoxin (Fd) and plastoquinone (PQ) within the membrane This pathway
di-verts the activated elost from PSI back through the PQ pool, the cytochrome b6f
complex, and plastocyanin to re-reduce P700(Figure 21.21)
Lumen
H2O
PQ
2 H+
4.5 H+
4.5 H+
4 H+
Cyt b6
Cyt b6 Cyt f
O2+ 2 H +
Q
QH 2
Plastocyanin docking
CF1CF 0 – ATP synthase
Stroma
H++ NADP+
FNR
NADPH
ADP + P i
Cu +
Cu 2+
PC
PC
Ferredoxin docking
Ferredoxin Fd
ATP
FeS A FeS B
FeS X
P700
A1
O O
O O
A0
Fe
PheD1 PheD2
ChlD1
ChlD2
PD1 PD2
D
Z-D2
(Mn)4
Photon
1
Photon
FIGURE 21.20 The mechanism of photophosphorylation Photosynthetic electron transport establishes a proton gradient that is tapped by the CF 1 CF 0 –ATP synthase to drive ATP synthesis.
Trang 8650 Chapter 21 Photosynthesis
Proton translocations accompany these cyclic electron transfer events so that ATP synthesis can be achieved In cyclic photophosphorylation, ATP is the sole product
of energy conversion No NADPH is generated, and because PSII is not involved, no oxygen is evolved Cyclic photophosphorylation theoretically yields 2 H per e
(2 H/h ) from the operation of the cytochrome b6f complex Thus, cyclic
photo-phosphorylation provides a mechanism for overcoming the ATP deficit for CO2 fix-ation (see the earlier section titled Calculfix-ation of the Photosynthetic Energy Re-quirements for Hexose Synthesis Depends on H/h and ATP/H Ratios, page 647) Estimates indicate that cyclic photophosphorylation may contribute about 10%
of total chloroplast ATP synthesis and thereby elevate the ATP/NADPH ratio
Organic Molecules?
As we began this chapter, we saw that photosynthesis traditionally is equated with the process of CO2fixation, that is, the net synthesis of carbohydrate from CO2 Indeed, the capacity to perform net accumulation of carbohydrate from CO2distinguishes the phototrophic (and autotrophic) organisms from heterotrophs Although animals possess enzymes capable of linking CO2to organic acceptors, they cannot achieve a net accumulation of organic material by these reactions For example, fatty acid biosynthesis is primed by covalent attachment of CO2to acetyl-CoA to form malonyl-CoA (see Chapter 24) Nevertheless, this “fixed CO2” is liberated in the very next re-action, so no net CO2incorporation occurs
Elucidation of the pathway of CO2fixation represents one of the earliest appli-cations of radioisotope tracers to the study of biology In 1945, Melvin Calvin and his colleagues at the University of California, Berkeley, were investigating photo-synthetic CO2 fixation in Chlorella Using14CO2, they traced the incorporation of radioactive14C into organic products and found that the earliest labeled product
was 3-phosphoglycerate (see Figure 17.14) Although this result suggested that the
CO2acceptor was a two-carbon compound, further investigation revealed that, in
ATP
ADP
H+
H+
Photon
Plastocyanin docking
Lumen Stroma
+ Pi
CF1
CF0
PC
H+
H+
FeS A FeS B
FeSX
P700
A1 O
O
O O
A0
PQ
Cyt b6
Cyt b6 Cyt f
FIGURE 21.21 The pathway of cyclic
photophosphoryla-tion by PSI (Adapted from Arnon, D I., 1984 The discovery of
photosynthetic phosphorylation Trends in Biochemical Sciences
9:258–262.)
Trang 9reality, 2 equivalents of 3-phosphoglycerate were formed following addition of CO2
to a five-carbon (pentose) sugar:
CO2 5-carbon acceptor ⎯⎯→ [6-carbon intermediate] ⎯⎯→
two 3-phosphoglycerates
Ribulose-1,5-Bisphosphate Is the CO2Acceptor in CO2Fixation
The five-carbon CO2 acceptor was identified as ribulose-1,5-bisphosphate (RuBP),
and the enzyme catalyzing this key reaction of CO2fixation is ribulose bisphosphate
carboxylase/oxygenase, or, in the jargon used by workers in this field, rubisco The
name ribulose bisphosphate carboxylase/oxygenase reflects the fact that rubisco catalyzes the
reaction of either CO2or, alternatively, O2with RuBP Rubisco is found in the
chloro-plast stroma It is a very abundant enzyme, constituting more than 15% of the total
chloroplast protein Given the preponderance of plant material in the biosphere,
rubisco is probably the world’s most abundant protein Rubisco is large: In higher
plants, rubisco is a 550-kD heteromultimeric (88) complex consisting of eight
iden-tical large subunits (55 kD) and eight small subunits (15 kD) (Figure 21.22) The large
subunit is the catalytic unit of the enzyme It binds both substrates (CO2and RuBP)
and Mg2(a divalent cation essential for enzymatic activity) The small subunit
modu-lates the catalytic efficiency of the enzyme, increasing kcatmore than 100-fold The
ru-bisco large subunit is encoded by a gene within the chloroplast DNA, whereas the
small subunit is encoded by a multigene family in the nuclear DNA Assembly of
ac-tive rubisco heteromultimers occurs within chloroplasts following transit of the
small subunit polypeptide across the chloroplast membrane
2-Carboxy-3-Keto-Arabinitol Is an Intermediate
in the Ribulose-1,5-Bisphosphate Carboxylase Reaction
The addition of CO2 to ribulose-1,5-bisphosphate results in the formation of an
enzyme-bound intermediate, 2-carboxy-3-keto-arabinitol (Figure 21.23, II) This
in-termediate arises when CO2 adds to the enediol intermediate generated from
ribulose-1,5-bisphosphate Hydrolysis of the C2OC3bond of the intermediate
gen-erates two molecules of 3-phosphoglycerate The CO2ends up as the carboxyl group
of one of the two molecules
Ribulose-1,5-Bisphosphate Carboxylase Exists in Inactive
and Active Forms
Rubisco exists in three forms: an inactive form, designated E; a carbamylated, but
in-active, form, designated EC; and an active form, ECM, which is carbamylated and has
Mg2at its active sites as well Carbamylation of rubisco takes place by addition of CO2
FIGURE 21.22 Molecular graphic of ribulose bisphos-phate carboxylase The enzyme consists of 8 equivalents each of two subunits Clusters of four small subunits (orange and red) are located at each end of the
sym-metric octamer formed by the L subunits (light and dark
green) The active sites are revealed in the ribbon dia-gram by bound ribulose-1,5-bisphosphate (yellow) (pdb id 1RXO).
H2COPO32– H2COPO32–
2
3
4
5
1
2 3 4 5
1
HCOH
HCOH
C COH HCOH
HCOH
HO
O
O–
HCOH C HO
III
O
O–
HCOH
C 2
H2O
CO2
FIGURE 21.23 The ribulose bisphosphate carboxylase reaction Mg 2 at the active site aids in stabilizing the
2,3-enediol transition state (I) for CO2 addition and in facilitating the carbon–carbon bond cleavage that leads to
product formation.
Trang 10652 Chapter 21 Photosynthesis
to its Lys201
2groups (to give derivatives) The CO2molecules used to carbamylate Lys residues do not become substrates The carbamylation reac-tion occurs spontaneously at slightly alkaline pH (pH 8) Carbamylareac-tion of rubisco completes the formation of a binding site for the Mg2that participates in the catalytic reaction Once Mg2 binds to EC, rubisco achieves its active ECM form Activated rubisco displays a K mfor CO2of 10 to 20 2in the atmosphere is low, about 0.03% The concentration of CO2dissolved in aqueous so-lutions equilibrated with air is about 10
Substrate RuBP binds much more tightly to the inactive E form of rubisco (KD
20 nM) than to the active ECM form (K mfor RuBP potent inhibitor of rubisco activity Release of RuBP from the active site of rubisco is
mediated by rubisco activase Rubisco activase is a regulatory protein; it binds to E-form
rubisco and, in an ATP-dependent reaction, promotes the release of RuBP Rubisco then becomes activated by carbamylation and Mg2binding Rubisco activase itself
is activated in an indirect manner by light Thus, light is the ultimate activator of rubisco
CO2Fixation into Carbohydrate Proceeds Via the Calvin–Benson Cycle
The immediate product of CO2fixation, 3-phosphoglycerate, must undergo a series
of transformations before the net synthesis of carbohydrate is realized Among car-bohydrates, hexoses (particularly glucose) occupy center stage Glucose is the build-ing block for both cellulose and starch synthesis These plant polymers constitute the most abundant organic material in the living world, and thus, the central focus on glucose as the ultimate end product of CO2fixation is amply justified Also, sucrose (-D-glucopyranosyl-(1⎯→2)--D-fructofuranoside) is the major carbon form translo-cated out of leaves to other plant tissues In nonphotosynthetic tissues, sucrose is me-tabolized via glycolysis and the TCA cycle to produce ATP
The set of reactions that transforms 3-phosphoglycerate into hexose is named
the Calvin–Benson cycle (often referred to simply as the Calvin cycle) for its
dis-coverers The reaction series is indeed cyclic because not only must carbohydrate appear as an end product, but the five-carbon acceptor, RuBP, must be regenerated
to provide for continual CO2fixation Balanced equations that schematically repre-sent this situation are
6(1) 6(5) ⎯⎯→ 12(3) 12(3)⎯⎯→ 1(6) 6(5)
Net: 6(1)⎯⎯→ 1(6) Each number in parentheses represents the number of carbon atoms in a com-pound, and the number preceding the parentheses indicates the stoichiometry
of the reaction Thus, 6(1), or 6 CO2, condense with 6(5) or 6 RuBP to give
12 3-phosphoglycerates These 12(3)s are then rearranged in the Calvin cycle to form one hexose, 1(6), and regenerate the six 5-carbon (RuBP) acceptors
The Enzymes of the Calvin Cycle Serve Three Metabolic Purposes
The Calvin cycle enzymes serve three important ends:
1 They constitute the predominant CO2fixation pathway in nature
2 They accomplish the reduction of 3-phosphoglycerate, the primary product of
CO2fixation, to glyceraldehyde-3-phosphate so that carbohydrate synthesis be-comes feasible
3 They catalyze reactions that transform three-carbon compounds into four-, five-, six-, and seven-carbon compounds
Most of the enzymes mediating the reactions of the Calvin cycle also participate
in either glycolysis (see Chapter 18) or the pentose phosphate pathway (see Chap-ter 22) The aim of the Calvin scheme is to account for hexose formation from