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ARS Home » Research » Publications at this Location » Publication #76595


item Crafts-Brandner, Steven
item Salvucci, Michael

Submitted to: Journal of Plant Physiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/1/1997
Publication Date: N/A
Citation: N/A

Interpretive Summary: The southwest cotton crop is often exposed to limited water and extremely high temperatures that can inhibit photosynthesis, thereby decreasing yield and quality. For cotton and other important crops, the heat-induced inhibition of photosynthesis is directly related to an effect on a chloroplast-localized enzyme called Rubisco activase. Rubisco activase functions to activate the enzyme, called Rubisco, that catalyzes the fixation of CO2 into sugars. Rubisco activase is found in two nearly identical forms in most plant species, but there is no information about the biological significance of this fact. We have used the cloned forms of Rubisco activase from spinach to determine the response of the two forms to high temperature. Our results demonstrate that the long form is much more tolerant of high temperatures than the short form. Significantly, when the two forms of the enzyme are mixed together the short form is protected against heat-inhibition. Our results provide the first scientific basis to form a hypothesis about the need for two forms of the enzyme. We will pursue research to determine how heat stress alters the metabolism of the two forms of Rubisco activase in cotton and we will determine if genetic differences in the production of the two forms in different crop species is related to high-temperature tolerance. This work is applicable to scientists in both the public and private sectors that are interested in plant response to environmental stress.

Technical Abstract: Ribulose-1,5-bisphosphate carboxylase/oxygenase activase often consists of two polypeptides that arise from alternative splicing of pre-mRNA. In this study, recombinant versions of the spinach (Spinacea oleracea L.) 45 kD and 41 kD forms of activase were analyzed for their response to temperature. The temperature optimum for ATPase hydrolysis by the 45 kD form was 45 deg. .C, approximately 13 deg. C higher than the 41 kD form. When the two forms were mixed, the temperature response of the hybrid enzyme was similar to the 45 kD form. In the absence of adenine nucleotide, preincubation of either activase form at temperatures above 25 deg. C inactivated ATPase activity (T50 = 30 deg. C). ATPgS, but not ADP, significantly enhanced the thermostability of the 45 kD form (T50 = 55 deg. C), but was much less effective for the 41 kD form (T50 = 41 deg. C). Measurements of intrinsic fluorescence showed that the increase that is associated with ATPgS-induced dsubunit aggregation was lost at a much lower temperature for the 41 kD for than for either the 45 kD form or a mixture of the two forms. However, the two activase forms were equally susceptible to limited proteolysis after heat treatment. The results indicate that 1) the two forms of activase differ in thermal stability; 2) the 45 kD form confers increased thermal stability to the 41 kD form in the hybrid enzyme; and 3) a loss of subunit interactions, rather than enzyme denaturation, appears to be the initial cause of temperature inactivation of activase.