A mutation in the large subunit of tobacco Rubisco reduces the decline in activity during catalysis

Abstract

The conversion of codon 335 of the large subunit of tobacco Rubisco to encode valine instead of leucine has been shown previously to reduce the maximum CO2-fixing rate and the enzyme¿s ability to distinguish between CO2 and O2 as substrates. Further studies show that the decline in activity during catalysis in vitro (fallover) is much less pronounced with the mutant enzyme. Both mutant and wild-type enzymes bind d-glycero-pentodiulose-1,5-bisphosphate (pentodiulose-P2), the dicarbonyl oxidation product of the substrate, d-ribulose-1,5-bisphosphate, resulting in a decrease in activity that can be reversed by the addition of Rubisco activase and ATP. The mutant enzyme is able to convert pentodiulose-P2 to 2-carboxytetritol-1,4-bisphosphate more rapidly and more completely than the wild-type enzyme. Both mutant and wild-type enzymes tightly bind 2¿-carboxy-d-arabinitol-1,5-bisphosphate (an analog of the six-carbon reaction intermediate) with complete abolition of catalytic activity. However, in both cases, the analog is released and carboxylation activity is restored by the action of Rubisco activase.