Possible explanation of the disparity between the in vitro and in vivo measurements of Rubisco activity: a study in loblolly pine grown in elevated pCO2

Abstract

Rubisco activity can be measured using gas exchange (in vivo) or using in vitro methods. Commonly in vitro methods yield activities that are less than those obtained in vivo. Rubisco activity was measured both in vivo and in vitro using a spectrophotometric technique in mature Pinus taeda L. (loblolly pine) trees grown using free-air CO2 enrichment in elevated (56 Pa) and current (36 Pa) pCO2. In addition, for studies where both in vivo and in vitro values of Rubisco activity were reported net CO2 uptake rate (A) was modelled based on the in vivo and in vitro values of Rubisco activity reported in the literature. Both the modelling exercise and the experimental data showed that the in vitro values of Rubisco activity were insufficient to account for the observed values of A. A trichloroacetic acid (TCA) precipitation of the protein from samples taken in parallel with those used for activity analysis was co-electrophoresed with the extract used for determining in vitro Rubisco activity. There was significantly more Rubisco present in the TCA precipitated samples, suggesting that the underestimation of Rubisco activity in vitro was attributable to an insufficient extraction of Rubisco protein prior to activity analysis. Correction of in vitro values to account for the under-represented Rubisco yielded mechanistically valid values for Rubisco activity. However, despite the low absolute values for Rubisco activity determined in vitro, the trends reported with CO2 treatment concurred with, and were of equal magnitude to, those observed in Rubisco activity measured in vivo.

A test of the ‘one‐point method’ for estimating maximum carboxylation capacity from field‐measured, light‐saturated photosynthesis

“…A closer match to in vitro enzyme activity of Rubisco can be obtained by considering the mesophyll conductance to CO 2 to the site of carboxylation (Rogers et al, 2001;Flexas et al, 2007); however, as g m values are available for so few of the sampled species, we assumed that C i is equal to C c , the CO 2 concentration at the chloroplast. The C i at which photosynthesis is colimited by both carboxylation and RuBP regeneration was calculated for each A-C i curve based on the apparent V cmax , J max and R day using the C 3 photosynthesis model.…”

Effects of water stress on photosynthesis and nitrogen metabolism in vegetative and reproductive shoots of Leymus chinensis

“…Besides, the great problem in RuBPCO activity assay in vitro is a low solubility of RuBPCO enzyme. Rogers et al (2001) showed that only 33 % of RuBPCO got into supernatant (or into filtration) during the extraction from the leaf. That is another reason why values V Cmax that express RuBPCO activity in vivo are higher than values of RuBPCO activity obtained in vitro; e.g.…”

Effect of water stress on dry mass accumulation and allocation in sweet potato based on stable isotope analysis

“…Then the mixture was centrifuged at 15 000 g for 10 min at 48C. The supernatant which was used to analyze RuBisCo activity was collected and measured by spectrophotometer at 340 nm using UV-1601 spectrophotometer (Rogers et al 2001). All statistical analyses were performed using trend analysis by SPSS v.11.5 (SPSS Inc., Chicago, IL).…”