Potential for interactions between the carboxy- and amino-termini of Rubisco activase subunits

The subunit interactions of Rubisco activase were investigated using mutants containing an introduced Cys near the N- and/or C-terminus. Chemical cross-linking of the C-terminal and double insertion mutant produced subunit dimers and dimers plus high ordered oligomers, respectively. Fluorescence measurements with N,N'-dimethyl-N-(iodoacetyl)-N'-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)ethylenediamine showed that the environment around the introduced Cys near the C-terminus becomes more hydrophilic upon nucleotide binding. The Cys insertion mutants catalyzed Rubisco activation and ATP hydrolysis even when the subunits of the C-terminal or double insertion mutants were completely cross-linked. The results indicate that the termini of adjacent activase subunits are in close proximity and can be modified and even joined without affecting enzyme function.