Facultad de Ciencias Exactas, Físicas y Naturales, Instituto Multidisciplinario de Biología Vegetal, CONICET, Universidad Nacional de Córdoba, Edificio de Investigaciones Biológicas y Tecnológicas, 5000 Córdoba, Argentina.
J Plant Res. 2012 Sep;125(5):653-60. doi: 10.1007/s10265-012-0476-8. Epub 2012 Jan 21.
Expression of the gene encoding the maize glycine-rich RNA-binding protein MA16 is developmentally regulated and it is involved in environmental stress responses. The MA16 protein shows a wide spectrum of RNA-binding activities. On the basis of in vivo labelling, where a [³²P]phosphate label was linked to the MA16 protein, Freire and Pages (Plant Mol Biol 29:797-807, 1995) suggested that the protein may be post-translationally modified by phosphorylation. However, further analysis showed that the [³²P]phosphate label was sensitive to different treatments, suggesting that modification distinct from protein phosphorylation might occur in the MA16 protein. Biochemical analysis revealed that this [³²P]phosphate labelling was resistant to phenol extraction and denaturing SDS-PAGE but sensitive to micrococcal nuclease, RNase A and RNase T1 treatments. The mobility of [³⁵S] labelled MA16 protein on SDS-PAGE did not significantly changed after the nuclease treatments suggesting that the [³²P]phosphate label associated to MA16 protein could be a ribonucleotide or a very short ribonucleotide chain. In addition, immunoprecipitation of labelled extracts showed that the ribonucleotide(s) linked to the MA16 protein was removed by phosphorolytic activity. This activity could be catalysed by a phosphate-dependent ribonuclease. The C-terminus of MA16 protein harbouring a glycine-rich domain was predicted to be an intrinsically disordered region.
编码玉米甘氨酸丰富的 RNA 结合蛋白 MA16 的基因的表达受到发育调控，并且它参与环境胁迫反应。MA16 蛋白表现出广泛的 RNA 结合活性。基于体内标记实验，其中[³²P]磷酸盐标记与 MA16 蛋白相连，Freire 和 Pages（植物分子生物学 29:797-807，1995）提出该蛋白可能通过磷酸化进行翻译后修饰。然而，进一步的分析表明，[³²P]磷酸盐标记对不同的处理敏感，表明 MA16 蛋白中可能发生不同于蛋白质磷酸化的修饰。生化分析表明，这种[³²P]磷酸盐标记对酚提取和变性 SDS-PAGE 具有抗性，但对微球菌核酸酶、RNase A 和 RNase T1 处理敏感。在核酶处理后，[³⁵S]标记的 MA16 蛋白在 SDS-PAGE 上的迁移率没有明显变化，这表明与 MA16 蛋白相关的[³²P]磷酸盐标记可能是核苷酸或非常短的核苷酸链。此外，标记提取物的免疫沉淀表明，与 MA16 蛋白相连的核苷酸被磷酸解活性去除。这种活性可以由依赖于磷酸盐的核糖核酸酶催化。MA16 蛋白的 C 末端含有一个甘氨酸丰富的结构域，被预测为一个固有无序区域。
