Proline caused irreversible inhibition (involving reduction in V-max without altering K-m for RuBP) in Rubisco activity. Proline-induced suppression in Rubisco activity did not exceed beyond similar to 65% of the original activity even upon exposure to higher levels of proline for prolonged duration. However, NaCl-induced reduction in Rubisco activity was reversible. Native PAGE analysis of Rubisco-incubated with proline showed the presence of two distinct bands corresponding to similar to 430 and similar to 28 kDa, but that incubated with NaCl showed a single band. SDS-PAGE analysis revealed that the similar to 430- and similar to 28-kDa bands represent octamers of large subunits and dimers of small subunits, respectively. These results demonstrated for the first time that proline suppresses Rubisco activity by bringing about dissociation of the small subunits from the octamer core of large subunits, probably by weakening hydrophobic interactions between them.