2nd net is up. Early bud formations are promising.  holding up to the extremes pretty well, some leaves taking minor damage, but overall, she is holding up, gave her 1 night at 50F see how she would react, stressful. Not advised as it messes with her metabolism, but I want to see if it triggers any anthocyanin response. Love to see her purp up but no signs yet.
Homework.
If Rubisco activity is impaired and it cannot properly function or regenerate its substrate, the plant's leaves are likely to turn a pale green or
lime green, a condition known as chlorosis. Essentially, Rubisco activity is highly regulated and susceptible to various environmental and metabolic factors that can cause it to become inhibited, leading to an apparent failure in RuBP regeneration due to a lack of consumption.
Rubisco regeneration is intrinsically linked to nitrogen supply because Rubisco is a major sink for nitrogen in plants, typically accounting for 15% to over 25% of total leaf nitrogen. The regeneration phase itself consumes nitrogen through the synthesis of the Rubisco enzyme and associated proteins (like Rubisco activase), and overall nitrogen status heavily influences the efficiency of RuBP regeneration. RuBisCO is a very large enzyme that constitutes a significant proportion (up to 50%) of leaf soluble protein and requires large investments in nitrogen. Insufficient nitrogen supply limits the plant's ability to produce adequate amounts of RuBisCO, thereby limiting the overall capacity for photosynthesis and carbon fixation. Maintaining the optimal, slightly alkaline pH is crucial for the proper function and regeneration of Rubisco. Deviations in either direction (too high or too low) disrupt the enzyme's structure, activation state, and interaction with its substrates, leading to decreased activity and impaired RuBP regeneration. (LIME GREEN CHLOROSIS)
Structural Component: Nitrogen is an essential building block for all proteins, and the sheer abundance of the Rubisco protein makes it the single largest storage of nitrogen in the leaf.
Synthesis and Activity: Adequate nitrogen supply is crucial for the synthesis and maintenance of sufficient Rubisco enzyme and Rubisco activase (Rca), the regulatory protein responsible for maintaining Rubisco's active state. Nitrogen deficiency leads to a decrease in the content and activity of both Rubisco and Rca, which in turn limits the maximum carboxylation rate, Vmax, and the rate of RuBP regeneration Jmax, thus reducing overall photosynthetic capacity.
Nitrogen Storage and Remobilization: Rubisco can act as a temporary nitrogen storage protein, which is degraded to remobilize nitrogen to other growing parts of the plant, especially under conditions of nitrogen deficiency or senescence.
Nitrogen Use Efficiency (NUE): The allocation of nitrogen to Rubisco is a key determinant of a plant's photosynthetic nitrogen use efficiency (PNUE). In high-nitrogen conditions, plants may accumulate a surplus of Rubisco, which may not be fully activated, leading to a lower PNUE. Optimizing the amount and activity of Rubisco relative to nitrogen availability is a target for improving crop NUE.
Photorespiration and Nitrogen Metabolism: Nitrogen metabolism is also linked to the photorespiration pathway (which competes with carboxylation at the Rubisco active site), particularly in the reassimilation of ammonia released during the process.
To increase RuBisCO regeneration, which refers to the process of forming the CO2 acceptor molecule Ribulose-1,5-bisphosphate (RuBP) during photosynthesis, the primary methods involve optimizing the levels and activity of Rubisco activase (Rca) and enhancing the performance of other Calvin-Benson-Bassham (CBB) cycle enzymes.
Biochemical and Environmental Approaches:
Optimize Rubisco Activase (Rca) activity: Rca is a crucial chaperone protein that removes inhibitory sugar phosphates, such as CA1P (2-carboxy-D-arabinitol 1-phosphate), from the Rubisco active site, thus maintaining its catalytic competence.
•Ensure optimal light conditions: Rca is light-activated via the chloroplast's redox status. Adequate light intensity ensures Rca can effectively maintain Rubisco in its active, carbamylated state.
•Maintain optimal temperature: Rca is highly temperature-sensitive and can become unstable at moderately high temperatures (e.g., above 35°C/95F° in many C3 plants), which decreases its ability to activate Rubisco. Maintaining temperatures within the optimal range for a specific plant species is important.
•Optimize Mg2+ concentration: Mg2+ is a key cofactor for both Rubisco carbamylation and Rca activity. In the light, Mg2+ concentration in the chloroplast stroma increases, promoting activation.
•Manage ATP/ADP ratio: Rca activity depends on ATP hydrolysis and is inhibited by ADP. Conditions that maintain a high ATP/ADP ratio in the chloroplast stroma favor Rca activity.
Enhance Calvin-Benson-Bassham (CBB) cycle enzyme activity: The overall rate of RuBP regeneration can be limited by other enzymes in the cycle.
•Increase SBPase activity: Sedoheptulose-1,7-bisphosphatase (SBPase) is a key regulatory enzyme in the regeneration pathway, and increasing its activity can enhance RuBP regeneration and overall photosynthesis.
•Optimize other enzymes: Overexpression of other CBB cycle enzymes such as fructose-1,6-bisphosphate aldolase (FBA) and triose phosphate isomerase (TPI) can also help to balance the metabolic flux and improve RuBP regeneration capacity.
Magnesium ions, Mg2+, are specifically required for Rubisco activation because the cation plays a critical structural and chemical role in forming the active site: A specific lysine residue in the active site must be carbamylated by a CO2 molecule to activate the enzyme. The resulting negatively charged carbamyl group then facilitates the binding of the positively charged Mg2+ion. While other divalent metal ions like Mn2+ can bind to Rubisco, they alter the enzyme's substrate specificity and lead to dramatically lower activity or a higher rate of the non-productive oxygenation reaction compared to Mg2+, making them biologically unfavorable in the context of efficient carbon fixation. The concentration of Mg2+ in the chloroplast stroma naturally increases in the light due to ion potential balancing during ATP synthesis, providing a physiological mechanism to ensure the enzyme is activated when photosynthesis is possible. At the center of the porphyrin ring, nestled within its nitrogen atoms, is a Magnesium ion (Mg2+). This magnesium ion is crucial for the function of chlorophyll, and without it, the pigment cannot effectively capture and transfer light energy.
Mg acts as a cofactor: Mg2+ binds to Rubisco after an activator CO2 molecule, forming a catalytically competent complex (Enzyme-CO2-Mg2+). High light + CO2) increases demand: Under high light (60 DLI is a very high intensity, potentially saturating) and high CO2, the plant's capacity for photosynthesis is high, and thus the demand for activated Rubisco and the necessary Mg2+ cofactor increases. Mg deficiency becomes limiting: If Mg2+ is deficient under these conditions, the higher levels of Rubisco and Rubisco activase produced cannot be fully activated, leading to lower photosynthetic rates and potential photo-oxidative damage.
Optimal range: Studies show that adequate Mg2+ application can enhance Rubisco activation and stabilize net photosynthetic rates under stress conditions, but the required concentration is specific to the experimental setup.
Monitoring is key: The most effective approach in a controlled environment is to monitor the plant's physiological responses e.g., leaf Mg2+ concentration, photosynthetic rate, Rubisco activation state, and adjust the nutrient solution/fertilizer to maintain adequate levels, rather than supplementing a fixed "extra" amount.
In practice, this means ensuring that Mg2+ is not a limiting factor in the plant's standard nutrient solution when pushing the limits with high light and CO2.
Applying Mg2+ through foliar spray is beneficial to Rubisco regeneration, particularly in alleviating the negative effects of magnesium (Mg) deficiency and high-temperature stress (HTS). While Mg can be leached from soil, within the plant it is considered a mobile nutrient, particularly in the phloem. Foliar-applied Mg is quickly absorbed by the leaves and can be translocate to other plant parts, including new growth and sink organs.
Foliar application of: NATURES VERY OWN MgSO4 @ 15.0g L-1 in a spray bottle.
Foliar sprays are often recommended as a rapid rescue measure for existing deficiencies or as a supplement during critical growth stages, when demand for Mg is high. Application in the early morning or late evening can improve absorption and prevent leaf burn.
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