High temperature enhances inhibitor production but reduces fallover in tobacco Rubisco

High temperature inhibits photosynthesis by several mechanisms including reduction in Rubisco activity. While the initial reaction velocity of purified, fully carbamylated, inhibitor-free Rubisco increases with temperature in vitro, over time, the reaction velocity slowly declines (fallover) because of the enzymatic and non-enzymatic production of inhibitors from the substrate ribulose-1,5-bisphosphate. We tested whether fallover could contribute to the decline in Rubisco activity observed in leaf extracts at high temperature. Production of d-xylulose-1,5-bisphosphate (XuBP), an inhibitor of Rubisco, was greater at 35 and 45°C than at 25°C but fallover was less severe at 35 and 45°C than at 25°C, both in rate and extent under saturating CO₂ and ambient O₂. This apparent dichotomy is consistent with the catalytic site of Rubisco loosening at higher temperatures and releasing inhibitors more easily. The loosening of the catalytic site was supported by the observation that RuBP and XuBP were released from their complexes with uncarbamylated, Mg²⁺-free Rubisco faster at 35 and 45°C than at 25°C. We conclude that, although XuBP production increased relative to catalytic throughput at higher temperatures, this was more than compensated for by its faster release, resulting in less fallover inhibition at higher temperatures.