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.
Salvucci
Michael E
ME
US Department of Agriculture, Agricultural Research Service, Western Cotton Research Laboratory, 4135 E. Broadway Road, Phoenix, AZ 85040-8803, USA. msalvucci@wcrl.ars.usda.gov
D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001483:Base Sequence; D002384:Catalysis; D003432:Cross-Linking Reagents / Q000737:chemistry; D003545:Cysteine / Q000378:metabolism; D019281:Dimerization; D004789:Enzyme Activation; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D016254:Mutagenesis, Insertional; D010940:Plant Proteins / Q000737:chemistry*; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D013329:Structure-Activity Relationship
D003432:Cross-Linking Reagents; D010940:Plant Proteins; D021122:Protein Subunits; D011994:Recombinant Proteins; C054765:rca protein, plant; D000255:Adenosine Triphosphate; D003545:Cysteine