Rubisco Extraction and Purification from Diatoms.

This protocol describes a method to extract ribulose-1,5-bisphosphate carboxylase oxygenase (Rubisco) from diatoms (Bacillariophyta) to determine catalytic performance. This protocol has been adapted from use in cyanobacteria and higher plants (Andrews, 1988; Whitney and Sharwood, 2007). First part (steps A1-A3) of the extraction provides a crude extract of Rubisco that is sufficient for carboxylation assays to measure the Michaelis constant for CO2 (KC) and the catalytic turnover rate ( kcat c ). However, the further purification steps outlined (steps B1-B4) are needed for measurements of Rubisco CO2/O2 Specificity (SC/O, [ Kane et al., 1994 ]).