Isolation, Expression and Characterization of rbc L Gene from Ulva prolifera J. Agardh(Ulvophyceae, Chlorophyta)  

作　　者：SHAO Zhanru LI Wei GUO Hui DUAN Delin 

机构地区：[1]Institute of Oceanology,Chinese Academy of Sciences [2]Institute of General Botany and Plant Physiology,Friedrich Schiller University [3]University of Chinese Academy of Sciences

出　　处：《Journal of Ocean University of China》2015年第6期1087-1095,共9页中国海洋大学学报（英文版）

基　　金：financially supported by the Scientific and Technical Supporting Programs of China(2008 BAC49B01);State Ocean Administration Project(200805069)

摘　　要：Ulva prolifera is a typical green alga in subtidal areas and can grow tremendously fast. A highly efficient Rubisco enzyme which is encoded by Up Rbc L gene may contribute to the rapid growth. In this study, the full-length Up Rbc L open reading frame(ORF) was identified, which encoded a protein of 474 amino acids. Phylogenetic analysis of Up Rbc L sequences revealed that Chlorophyta had a closer genetic relationship with higher plants than with Rhodophyta and Phaeophyta. The two distinct residues(aa11 and aa91) were presumed to be unique for Rubisco catalytic activity. The predicted three-dimensional structure showed that one α/β-barrel existed in the C-terminal region, and the sites for Mg^(2+)coordination and CO_2 fixation were also located in this region. Gene expression profile indicated that Up Rbc L was expressed at a higher level under light exposure than in darkness. When the culture temperature reached 35℃, the expression level of Up Rbc L was 2.5-fold lower than at 15℃, and the carboxylase activity exhibited 13.8-fold decrease. Up Rbc L was heterologously expressed in E. coli and was purified by Ni^(2+) affinity chromatography. The physiological and biochemical characterization of recombinant Rubisco will be explored in the future.Ulva prolifera is a typical green alga in subtidal areas and can grow tremendously fast. A highly efficient Rubisco enzyme which is encoded by UpRbcL gene may contribute to the rapid growth. In this study, the full-length UpRbcL open reading frame (ORF) was identified, which encoded a protein of 474 amino acids. Phylogenetic analysis of UpRbcL sequences revealed that Chlorophyta had a closer genetic relationship with higher plants than with Rhodophyta and Phaeophyta. The two distinct residues (aa11 and aa91) were presumed to be unique for Rubisco catalytic activity. The predicted three-dimensional structure showed that one alpha/beta-barrel existed in the C-terminal region, and the sites for Mg2+ coordination and CO2 fixation were also located in this region. Gene expression profile indicated that UpRbcL was expressed at a higher level under light exposure than in darkness. When the culture temperature reached 35A degrees C, the expression level of UpRbcL was 2.5-fold lower than at 15A degrees C, and the carboxylase activity exhibited 13.8-fold decrease. UpRbcL was heterologously expressed in E. coli and was purified by Ni2+ affinity chromatography. The physiological and biochemical characterization of recombinant Rubisco will be explored in the future.

关 键 词：Ribulose-1 5-bisphosphate carboxylase/oxygenase large subunit sequence analysis real-time PCR in vitro expres-sion ULVA PROLIFERA 

分 类 号：S917.3[农业科学—水产科学]