Autotrophic Assimilation – Photosynthesis

Overview of Autotrophic Assimilation

“Autotrophic assimilation” in biology refers to the ability of certain organisms (autotrophs) to build organic compounds from inorganic starting materials. In most ecosystems, the most important autotrophic process is photosynthesis: the conversion of light energy into chemical energy stored in organic molecules, mainly carbohydrates.

In this chapter the focus is on:

• How light energy is converted into chemical energy (ATP, reduced cofactors)
• How inorganic carbon ($\mathrm{CO_2}$) is incorporated into organic molecules
• The main structures and organic reactions involved in photosynthesis
• How photosynthesis connects to the broader organic chemistry of living systems

General metabolism and biocatalysis are treated in the parent chapter; here we emphasize what is specific to photosynthetic autotrophic assimilation.

Photosynthetic Organisms and Cellular Structures

Photosynthetic Organisms

Organisms performing photosynthesis include:

• Plants (photosynthesis in chloroplasts of leaf cells)
• Algae (eukaryotic; chloroplasts similar to plants)
• Cyanobacteria (prokaryotic; photosynthetic membranes, but no chloroplasts)

All of them convert $\mathrm{CO_2}$ and water into organic matter, using light as the primary energy source and producing $\mathrm{O_2}$ (in oxygenic photosynthesis).

A simplified overall equation for oxygenic photosynthesis is:

$$
6\,\mathrm{CO_2} + 6\,\mathrm{H_2O} \xrightarrow{\text{light}} \mathrm{C_6H_{12}O_6} + 6\,\mathrm{O_2}
$$

Here $\mathrm{C_6H_{12}O_6}$ represents a hexose such as glucose; in reality, plants first form triose phosphates and then a variety of carbohydrates.

Chloroplast Structure (Plants and Algae)

In plants and algae, photosynthesis takes place in chloroplasts. Key structural features that are important for the chemistry:

• Outer and inner membrane: enclose the organelle.
• Stroma: aqueous phase inside the inner membrane, rich in enzymes, nucleotides, and inorganic ions. This is where the carbon fixation reactions occur.
• Thylakoid membranes: internal membrane system where the light-dependent reactions occur.
• Stacked regions form grana; these are interconnected by stroma lamellae.
• The interior space of the thylakoids is the thylakoid lumen.

The physical separation between stroma and thylakoid lumen is crucial for setting up and using proton gradients and for organizing electron transfer chains.

Cyanobacteria lack chloroplasts but have internal membrane systems functionally analogous to thylakoids.

Light-Dependent Reactions: Conversion of Light to Chemical Energy

The light-dependent reactions transform light energy into:

• A transmembrane proton gradient (electrochemical potential)
• ATP (from ADP and inorganic phosphate $\mathrm{P_i}$)
• A reduced cofactor (in plants: $\mathrm{NADPH}$)

These reactions are located in the thylakoid membrane and involve specialized pigment–protein complexes.

Photosynthetic Pigments and Light Absorption

The primary pigments in oxygenic photosynthesis are:

• Chlorophylls (mainly chlorophyll a, and also chlorophyll b in plants)
• Macrocyclic, porphyrin-like structure with a central $\mathrm{Mg^{2+}}$ ion
• Extended conjugated $\pi$ system; this delocalization allows absorption of visible light and excitation of electrons
• Accessory pigments:
• Carotenoids (polyene chains, strongly conjugated; absorb blue–green light, protect against photooxidative damage)
• Additional chlorophyll types or phycobiliproteins in some organisms

These pigments are organized into:

• Light-harvesting complexes (LHCs): arrays of pigments bound to proteins that capture photons and transfer excitation energy to reaction centers via resonance energy transfer.
• Reaction centers: specialized pigment–protein complexes where charge separation (conversion of excited states into separated charges) occurs.

From an organic-chemistry viewpoint, conjugated double-bond systems are key; excitation of electrons in these systems enables the primary photophysical and photochemical events.

Photosystems and Electron Transport

In plants and algae, two photosystems cooperate:

• Photosystem II (PSII): oxidizes water, releases $\mathrm{O_2}$, and starts the electron transport chain.
• Photosystem I (PSI): uses light energy to drive the final reduction of $\mathrm{NADP^+}$ to $\mathrm{NADPH}$.

Water Splitting and Oxygen Evolution

In PSII:

• Light absorption leads to an excited chlorophyll in the reaction center designated P680.
• P680* donates an electron to a series of acceptors; P680 becomes a strong oxidant P680$^+$.
• P680$^+$ extracts electrons from water via a manganese-calcium cluster (the oxygen-evolving complex, OEC).

Net reaction in water splitting:

$$
2\,\mathrm{H_2O} \rightarrow 4\,\mathrm{H^+} + 4\,\mathrm{e^-} + \mathrm{O_2}
$$

The protons are released into the thylakoid lumen, contributing to the proton gradient. Electrons are passed through plastoquinone and other carriers to cytochrome $b_6f$.

Electron Transport and Proton Gradient

Electrons travel from PSII to PSI through carriers including:

• Plastoquinone (PQ / PQH$_2$): a benzoquinone derivative with an isoprenoid side chain; shuttles electrons and protons in the thylakoid membrane.
• Cytochrome $b_6f$ complex: mediates electron transfer and pumps protons into the lumen.
• Plastocyanin: a soluble copper protein in the lumen; carries electrons to PSI.

At PSI:

• P700 chlorophyll absorbs light; excited P700* donates an electron down a chain of acceptors.
• Final electron acceptor in this chain is ferredoxin, a small iron–sulfur protein in the stroma.

The electron transfer chain has two major consequences:

1. Proton accumulation in the thylakoid lumen
• From water oxidation
• From PQ/PQH$_2$ cycling through cytochrome $b_6f$
2. Reduction of $\mathrm{NADP^+}$ in the stroma:
   $$
   \mathrm{NADP^+ + H^+ + 2\,e^- \rightarrow NADPH}
   $$

The resulting proton-motive force across the thylakoid membrane is used to synthesize ATP.

Photophosphorylation: ATP Synthesis

ATP synthase is embedded in the thylakoid membrane. It couples proton flow from the lumen back into the stroma to the endergonic synthesis of ATP:

$$
\mathrm{ADP + P_i \xrightarrow{\text{ATP synthase}} ATP}
$$

Because the driving force is light, this form of ATP synthesis in chloroplasts is termed photophosphorylation.

Key point: after the light reactions, the stroma contains:

• ATP (chemical energy)
• $\mathrm{NADPH}$ (reducing power)

Both are essential for the carbon fixation reactions of the Calvin cycle.

Carbon Fixation: The Calvin–Benson–Bassham Cycle

The Calvin cycle (often simply “Calvin cycle”) is the central pathway for autotrophic assimilation of $\mathrm{CO_2}$ in most plants and many photosynthetic microorganisms. It runs in the stroma of chloroplasts.

The core function:

• Incorporate inorganic $\mathrm{CO_2}$ into organic molecules
• Use ATP and $\mathrm{NADPH}$ from the light reactions to reduce and rearrange these carbon skeletons
• Regenerate the initial acceptor for $\mathrm{CO_2}$

The cycle can be divided into three phases:

1. Carboxylation
2. Reduction
3. Regeneration

Phase 1: Carboxylation (CO₂ Fixation)

The initial acceptor for $\mathrm{CO_2}$ is a 5-carbon sugar:

• Ribulose-1,5-bisphosphate (RuBP)

The key enzyme:

• Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco)

Rubisco catalyzes the carboxylation:

$$
\mathrm{RuBP (C_5) + CO_2 \rightarrow 2\; 3\text{-phosphoglycerate} \ (2 \times C_3)}
$$

In more detail, the 6-carbon intermediate formed after $\mathrm{CO_2}$ addition is unstable and splits into two molecules of 3-phosphoglycerate (3-PGA), each with three carbons.

From an organic perspective, this is a nucleophilic addition of $\mathrm{CO_2}$ to a sugar phosphate, followed by cleavage.

Phase 2: Reduction

The two 3-PGA molecules are then:

1. Phosphorylated by ATP:
   $$
   \mathrm{3\text{-PGA} + ATP \rightarrow 1,3\text{-bisphosphoglycerate} + ADP}
   $$
2. Reduced by $\mathrm{NADPH}$:
   $$
   \mathrm{1,3\text{-bisphosphoglycerate} + NADPH + H^+ \rightarrow glyceraldehyde\text{-}3\text{-phosphate} \ (G3P) + P_i + NADP^+}
   $$

The product glyceraldehyde-3-phosphate (G3P) is a triose phosphate.

Organic viewpoint:

• The carboxyl group in 3-PGA is first activated as a mixed acid anhydride (1,3-bisphosphoglycerate).
• Then it is reduced to an aldehyde (G3P) using $\mathrm{NADPH}$.
• This is analogous to steps in glycolysis but operates in reverse direction.

Phase 3: Regeneration of Ribulose-1,5-bisphosphate

Most of the G3P produced is not exported but used to regenerate RuBP. This regeneration involves multiple sugar-phosphate rearrangements:

• Interconversions between triose phosphates (C$_3$), tetrose phosphates (C$_4$), pentose phosphates (C$_5$), and hexose/septose intermediates (C$_6$, C$_7$).
• Enzymes such as:
• Transketolase (transfers a 2-carbon unit between sugar phosphates)
• Aldolase (condenses shorter sugars to longer ones)

At the end, ribulose-5-phosphate is formed and then phosphorylated by ATP:

$$
\mathrm{ribulose\text{-}5\text{-phosphate} + ATP \rightarrow ribulose\text{-}1,5\text{-bisphosphate} + ADP}
$$

This closes the cycle and prepares another RuBP molecule to accept $\mathrm{CO_2}$.

Stoichiometry of the Calvin Cycle

To produce one net triose phosphate (G3P) that can leave the cycle:

• 3 molecules of $\mathrm{CO_2}$ are fixed.
• 3 RuBP (C$_5$) + 3 $\mathrm{CO_2}$ → 6 3-PGA (C$_3$).
• 1 of the resulting G3P (C$_3$ equivalent) is net gain; the rest regenerate RuBP.

The overall balanced stoichiometry for synthesis of one G3P is:

$$
3\,\mathrm{CO_2} + 9\,\mathrm{ATP} + 6\,\mathrm{NADPH} + 5\,\mathrm{H_2O}
\rightarrow
\mathrm{G3P} + 9\,\mathrm{ADP} + 8\,\mathrm{P_i} + 6\,\mathrm{NADP^+} + 2\,\mathrm{H^+}
$$

Later, two G3P molecules can be combined to form hexoses like fructose-1,6-bisphosphate and eventually starch or sucrose.

Organic Products of Photosynthesis and Their Fates

The immediate organic product of the Calvin cycle is G3P (a triose phosphate). From this central intermediate, various classes of organic compounds are synthesized.

Carbohydrate Synthesis

From G3P, plants synthesize:

• Hexose phosphates:
• Fructose-6-phosphate
• Glucose-6-phosphate
• Glucose-1-phosphate

These are precursors for:

• Starch (storage polysaccharide in chloroplasts and amyloplasts)
• Sucrose (transport form of carbohydrate in many plants; synthesized mainly in the cytosol)
• Cellulose (structural polysaccharide in cell walls; synthesized at the plasma membrane from UDP-glucose)

Key organic reactions involved:

• Aldol condensations (e.g., combining two G3P to form fructose-1,6-bisphosphate)
• Isomerizations between aldoses and ketoses
• Phosphoryl transfer (kinase and phosphatase reactions)
• Formation of glycosidic bonds in polysaccharide biosynthesis

Precursors for Other Biomolecules

Because G3P can be further metabolized, photosynthesis indirectly provides carbon skeletons for:

• Amino acids: via intermediates of central metabolism (e.g., 3-PGA, pyruvate, oxaloacetate, and $\alpha$-ketoglutarate derived pathways).
• Lipids: glycerol backbone (from G3P derivatives) and fatty acids (from acetyl-CoA derived from pyruvate).
• Nucleotides: ribose-5-phosphate and other pentose phosphates come from the pentose phosphate pathways linked to Calvin cycle intermediates.

Thus, photosynthesis is the primary source of reduced carbon in most ecosystems, feeding into the synthesis of numerous organic functional groups and compounds discussed elsewhere in the course.

Variants of Carbon Fixation: C₃, C₄, and CAM Pathways (Overview)

The Calvin cycle itself produces a 3-carbon compound (3-PGA) as the first stable product of $\mathrm{CO_2}$ fixation; plants relying solely on this are called C$_3$ plants.

To reduce losses due to photorespiration (a side reaction of Rubisco with $\mathrm{O_2}$), certain plants evolved additional $\mathrm{CO_2}$-concentrating mechanisms:

• C$_4$ pathway:
• First $\mathrm{CO_2}$ is fixed into 4-carbon acids (e.g., oxaloacetate, malate) by the enzyme PEP carboxylase.
• These acids transport $\mathrm{CO_2}$ to specialized cells where the Calvin cycle operates at elevated $\mathrm{CO_2}$ concentration.
• CAM (Crassulacean Acid Metabolism):
• Stomata open at night; $\mathrm{CO_2}$ is fixed into organic acids (often malate) and stored in vacuoles.
• During the day, $\mathrm{CO_2}$ is released internally and used by the Calvin cycle.

Chemically, both strategies involve additional carboxylation and decarboxylation reactions and temporary storage of carbon as organic acids; the Calvin cycle remains the final stage of $\mathrm{CO_2}$ assimilation into carbohydrates.

Energetics and Redox Aspects

From the standpoint of thermodynamics and redox chemistry, photosynthesis performs:

• An overall reduction of carbon:
• Carbon in $\mathrm{CO_2}$ is in a high oxidation state.
• Carbon in carbohydrates is more reduced.
• A coupled oxidation of water:
• Water loses electrons and is converted to $\mathrm{O_2}$.

By convention, the overall process can be described as:

$$
\mathrm{CO_2} + \mathrm{H_2O} \xrightarrow{\text{light}} \text{“CH}_2\text{O”} + \mathrm{O_2}
$$

where “CH$_2$O” represents the average composition of carbohydrate.

Key redox agent:

• $\mathrm{NADP^+ / NADPH}$ pair, which mediates hydride transfer in many biosynthetic reductions, including the Calvin cycle.

The energy for these uphill transformations comes from absorbed light; chemically, this energy is first captured in excited electronic states and then stored in proton gradients, ATP, and reduced cofactors.

Ecological and Biogeochemical Significance

While this course focuses on molecular structures and reactions, it is important to note the broader consequences of photosynthetic autotrophic assimilation:

• It is the primary entry point of solar energy into the biosphere.
• It produces the bulk of atmospheric $\mathrm{O_2}$.
• It drives major global biogeochemical cycles:
• Carbon cycle (exchange between $\mathrm{CO_2}$ and organic carbon)
• Coupled cycles of oxygen, nitrogen, and others via biologically produced organic matter.

Photosynthesis thus links the organic chemistry of living systems to the composition of the atmosphere, hydrosphere, and lithosphere.

Summary

• Autotrophic assimilation by photosynthesis converts light energy, water, and $\mathrm{CO_2}$ into chemical energy and reduced carbon compounds.
• Light reactions in the thylakoid membrane use chlorophyll and other pigments to carry out water oxidation, electron transport, proton pumping, $\mathrm{NADP^+}$ reduction, and ATP synthesis.
• The Calvin cycle in the stroma fixes $\mathrm{CO_2}$ into 3-phosphoglycerate, reduces it to G3P using ATP and $\mathrm{NADPH}$, and regenerates ribulose-1,5-bisphosphate.
• The immediate product G3P serves as a central building block for carbohydrates and, via metabolic interconversions, for lipids, amino acids, and many other biomolecules.
• Variants such as C$_4$ and CAM metabolism modify the initial steps of $\mathrm{CO_2}$ fixation but ultimately rely on the Calvin cycle.
• Photosynthesis is the chemical foundation of most life on Earth, coupling inorganic and organic worlds through redox and energy-transforming reactions.