State Key Laboratory of Plant Molecular Genetics, CAS Center for Excellence in Molecular Plant Sciences, 200032 Shanghai, China; University of Chinese Academy of Sciences, 200032 Shanghai, China.
CAS Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, 200032 Shanghai, China; University of Chinese Academy of Sciences, 200032 Shanghai, China.
Mol Plant. 2016 Jul 6;9(7):1018-27. doi: 10.1016/j.molp.2016.03.012. Epub 2016 Apr 19.
Atmospheric carbon dioxide (CO2) is assimilated by the most abundant but sluggish enzyme, ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco). Here we show that acetylation of lysine residues of the Rubisco large subunit (RbcL), including Lys201 and Lys334 in the active sites, may be an important mechanism in the regulation of Rubisco activities. It is well known that Lys201 reacts with CO2 for carbamylation, a prerequisite for both carboxylase and oxygenase activities of Rubisco, and Lys334 contacts with ribulose-1,5-bisphosphate (RuBP). The acetylation level of RbcL in plants is lower during the day and higher at night, inversely correlating with the Rubisco carboxylation activity. A search of the chloroplast proteome database did not reveal a canonical acetyltransferase; instead, we found that a plant-derived metabolite, 7-acetoxy-4-methylcoumarin (AMC), can non-enzymatically acetylate both native Rubisco and synthesized RbcL peptides spanning Lys334 or Lys201. Furthermore, lysine residues were modified by synthesized 4-methylumbelliferone esters with different electro- and stereo-substitutes, resulting in varied Rubisco activities. 1-Chloroethyl 4-methylcoumarin-7-yl carbonate (ClMC) could transfer the chloroethyl carbamate group to lysine residues of RbcL and completely inactivate Rubisco, whereas bis(4-methylcoumarin-7-yl) carbonate (BMC) improved Rubisco activity through increasing the level of Lys201 carbamylation. Our findings indicate that RbcL acetylation negatively regulates Rubisco activity, and metabolic derivatives can be designed to dissect and improve CO2 fixation efficiency of plants through lysine modification.
大气中的二氧化碳（CO2）被最丰富但最缓慢的酶——核酮糖-1,5-二磷酸羧化酶/加氧酶（Rubisco）同化。在这里，我们表明 Rubisco 大亚基（RbcL）赖氨酸残基的乙酰化，包括活性部位的赖氨酸 201 和赖氨酸 334，可能是调节 Rubisco 活性的重要机制。众所周知，赖氨酸 201 与 CO2 反应发生氨甲酰化，这是 Rubisco 羧化酶和加氧酶活性的前提条件，而赖氨酸 334 与核酮糖-1,5-二磷酸（RuBP）结合。植物中 RbcL 的乙酰化水平在白天较低，在夜间较高，与 Rubisco 羧化活性呈负相关。在叶绿体蛋白质组数据库中搜索并没有发现典型的乙酰转移酶；相反，我们发现一种植物衍生的代谢物，7-乙酰氧基-4-甲基香豆素（AMC），可以非酶促地乙酰化天然 Rubisco 和合成的跨越赖氨酸 334 或赖氨酸 201 的 RbcL 肽。此外，赖氨酸残基被具有不同电性和立体取代基的合成 4-甲基伞形酮酯修饰，导致 Rubisco 活性不同。1-氯乙基 4-甲基香豆素-7-基碳酸酯（ClMC）可以将氯乙基氨基甲酸酯基团转移到 RbcL 的赖氨酸残基上，从而完全使 Rubisco 失活，而双（4-甲基香豆素-7-基）碳酸酯（BMC）通过增加赖氨酸 201 的氨甲酰化水平来提高 Rubisco 活性。我们的研究结果表明，RbcL 乙酰化负调节 Rubisco 活性，并且代谢衍生物可以通过赖氨酸修饰来设计以剖析和提高植物的 CO2 固定效率。
