Microtubule associated protein 2 or MAP2 is a major microtubule binding protein of neurons, where it is concentrated in dendrites and perikarya but absent from axons (1). There is a single mammalian MAP2 gene which may generate multiple lower molecular weight forms usually named MAP2C and MAP2D which run on SDS-PAGE gels at 60-70kDa, though are actually much smaller in molecular size. These forms are found early in development but as the animal matures are replaced by MAP2A and B which are much larger in molecular size. These two forms include the so-called projection domain, a long insert which results in both molecules running at ~240kDa on SDS-PAGE gels. All four forms contain an N-terminal region which includes a binding site of cAMP dependent protein kinase (1). We have expressed five different recombinant forms of MAP2 based on the human sequence reported in REFSEQ XP_006712595.. These constructs in combination cover almost the entire human MAP2 molecule. We have used these constructs to generate a series of monoclonal and polyclonal antibodies to MAP2, many of which are known to recognize defined segments of MAP2.
We inserted cDNAs encoding the various MAP2 sequences into the pET29a eukaryotic expression vector. The vector adds an C-terminal His-tag which was use to purify the protein and this, along with some other vector derived sequence, adds about 5 kDa to the molecule.
The sequence is amino acids 712-1136 from REFSEQ XP_006712595. This corresponds to the second segment of the “projection domain” a long insert expressed in MAP2A and MAP2B but absent from MAP2C and MAP2D. This insert is responsible for thin side arms seen on microtubules in neuronal dendrites. The construct was expressed in pET29a and has a few N and C-terminal amino acids derived from the vector, including a C-terminal His-tag.
>MAP2-P2 SVHESIDTMSPMHKNGDKEFQTGKESQPSPPAQEAGYSTLAQSYPSDLPEEPSSPQERMF TIDPKVYGEKRDLHSKNKDDLTLSRSLGLGGRSAIEQRSMSINLPMSCLDSIALGFNFGR GHDLSPLASDILTNTSGSMDEGDDYLPATTPALEKAPCFPVESKEEEQIEKVKATGEEST QAEISCESPFLAKDFYKNGTVMAPDLPEMLDLAGTRSRLASVSADAEVARRKSVPSETVV EDSRTGLPPVTDENHVIVKTDSQLEDLGYCVFNKYTVPLPSPVQDSENLSGESGTFYEGT DDKVRRDLATDLSLIEVKLAAAGRVKDEFSVDKEASAHISGDKSGLSKEFDQEKKANDRL DTVLEKSEEHADSKEHAKKTEEAGDEIETFGLGVTYEQALAKDLSIPTDASSEKAEKGLS SVPEIAEVEPSKKVEQGLDFAVQGQLDVKISDFGQMASGLNIDDRRATELKLEATQDMTP
