Wang Dafu, Naidu Shawna L, Portis Archie R, Moose Stephen P, Long Stephen P
Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA.
J Exp Bot. 2008;59(7):1779-87. doi: 10.1093/jxb/ern074.
The previous investigations show that the amount and activity of Rubisco appears the major limitation to effective C(4) photosynthesis at low temperatures. The chilling-tolerant and bioenergy feedstock species Miscanthus x giganteus (M. x giganteus) is exceptionally productive among C(4) grasses in cold climates. It is able to develop photosynthetically active leaves at temperatures 6 degrees C below the minimum for maize, and achieves a productivity even at 52 degrees N that exceeds that of the most productive C(3) crops at this latitude. This study investigates whether this unusual low temperature tolerance can be attributed to differences in the amount or kinetic properties of Rubisco relative to maize. An efficient protocol was developed to purify large amounts of functional Rubisco from C(4) leaves. The maximum carboxylation activities (V(max)), activation states, catalytic rates per active site (K(cat)) and activation energies (E(a)) of purified Rubisco and Rubisco in crude leaf extracts were determined for M. x giganteus grown at 14 degrees C and 25 degrees C, and maize grown at 25 degrees C. The sequences of M. x giganteus Rubisco small subunit mRNA are highly conserved, and 91% identical to those of maize. Although there were a few differences between the species in the translated protein sequences, there were no significant differences in the catalytic properties (V(max), K(cat), and E(a)) for purified Rubisco, nor was there any effect of growth temperature in M. x giganteus on these kinetic properties. Extracted activities were close to the observed rates of CO(2) assimilation by the leaves in vivo. On a leaf area basis the extracted activities and activation state of Rubisco did not differ significantly, either between the two species or between growth temperatures. The activation state of Rubisco in leaf extracts showed no significant difference between warm and cold-grown M. x giganteus. In total, these results suggest that the ability of M. x giganteus to be productive and maintain photosynthetically competent leaves at low temperature does not result from low temperature acclimation or adaptation of the catalytic properties of Rubisco.
先前的研究表明，核酮糖-1,5-二磷酸羧化酶（Rubisco）的含量和活性似乎是低温下C4光合作用效率的主要限制因素。耐寒且作为生物能源原料的芒草（Miscanthus x giganteus，简称M. x giganteus）在寒冷气候下的C4禾本科植物中具有极高的生产力。它能够在比玉米最低温度低6摄氏度的温度下长出具有光合活性的叶片，甚至在北纬52度的地区也能实现超过该纬度最具生产力的C3作物的产量。本研究调查了这种异常的低温耐受性是否可归因于与玉米相比，Rubisco在含量或动力学特性上的差异。我们开发了一种有效的方法，从C4叶片中纯化出大量有功能的Rubisco。测定了在14摄氏度和25摄氏度下生长的M. x giganteus以及在25摄氏度下生长的玉米的纯化Rubisco和粗叶提取物中Rubisco的最大羧化活性（V(max)）、活化状态、每个活性位点的催化速率（K(cat)）和活化能（E(a)）。M. x giganteus Rubisco小亚基mRNA的序列高度保守，与玉米的序列有91%的同一性。尽管在翻译后的蛋白质序列中，不同物种之间存在一些差异，但纯化的Rubisco在催化特性（V(max)、K(cat)和E(a)）方面没有显著差异，生长温度对M. x giganteus的这些动力学特性也没有任何影响。提取的活性接近叶片在体内观察到的CO2同化速率。以叶面积为基础，Rubisco的提取活性和活化状态在两个物种之间或生长温度之间均无显著差异。在温暖和寒冷环境下生长的M. x giganteus叶片提取物中，Rubisco的活化状态没有显著差异。总体而言，这些结果表明，M. x giganteus在低温下保持高产并维持具有光合能力叶片的能力并非源于Rubisco催化特性的低温驯化或适应。
