Sánchez C, Tompa P, Szücs K, Friedrich P, Avila J
Centro de Biología Molecular Severo Ochoa, Facultad de Ciencias, Universidad Autónoma de Madrid, Spain.
Eur J Biochem. 1996 Nov 1;241(3):765-71. doi: 10.1111/j.1432-1033.1996.00765.x.
The C-terminal domain of microtubule-associated protein 2 (MAP2) contains a proline-rich region and the tubulin-binding domain. We have generated antibodies to follow the phosphorylation state of the proline-rich domain. One of these antibodies (no. 305) has been raised against a synthetic peptide P (sequence RTPGTPGTPSY) phosphorylated at the threonine residues. This sequence is present in the proline-rich region of MAP2 and is phosphorylated in vitro by at least three different proline-directed protein kinases: p42mpk, p34cdc2, and GSK3 (glycogen-synthase kinase 3) alpha/beta. The MAP2 sites phosphorylated by these kinases are different, although all of them phosphorylate the C-terminal domain of MAP2 as determined by Staphylococcus aureus V8 protease mapping. Nonphosphorylated peptide P can be phosphorylated in vitro by all three kinases studied with similar efficiency. In high-molecular-mass MAP2, this sequence is highly phosphorylated in vivo at the late stages of rat development. This motif can be rapidly dephosphorylated in vitro by protein-phosphatase 1 (PP1) and 2A (PP2A) catalytic subunits but not by PP2B.
微管相关蛋白2（MAP2）的C末端结构域包含一个富含脯氨酸的区域和微管蛋白结合结构域。我们制备了抗体来追踪富含脯氨酸结构域的磷酸化状态。其中一种抗体（编号305）是针对在苏氨酸残基处磷酸化的合成肽P（序列为RTPGTPGTPSY）产生的。该序列存在于MAP2的富含脯氨酸区域，并且在体外可被至少三种不同的脯氨酸定向蛋白激酶磷酸化：p42mpk、p34cdc2和糖原合酶激酶3（GSK3）α/β。尽管通过金黄色葡萄球菌V8蛋白酶图谱分析确定这些激酶磷酸化的MAP2位点不同，但它们均磷酸化MAP2的C末端结构域。未磷酸化的肽P在体外可被所研究的所有三种激酶以相似的效率磷酸化。在高分子量的MAP2中，该序列在大鼠发育后期在体内高度磷酸化。该基序在体外可被蛋白磷酸酶1（PP1）和2A（PP2A）催化亚基快速去磷酸化，但不能被PP2B去磷酸化。
