PRMT3 is a distinct member of the protein arginine N-methyltransferase family - Conferral of substrate specificity by a zinc-finger domain

S-Adenosyl-L-methionine-dependent protein arginine N-methyltransferases (PR MTs) catalyze the methylation of arginine residues within a variety of prot eins. At least four distinct mammalian family members have now been describ ed, including PRMT1, PRMT3, CARM1/PRMT4, and JBP1/PRMT5. To more fully defi ne the physiological role of PRMT3, we characterized its unique putative zi nc-finger domain and how it can affect its enzymatic activity. Here we show that PRMT3 does contain a single zinc-finger domain in its amino terminus. Although the zinc-liganded form of this domain is not required for methyla tion of an artificial substrate such as the glutathione S-transferase-fibri llarin amino-terminal fusion protein (GST-GAR), it is required for the enzy me to recognize RNA-associated substrates in RAT1 cell extracts. The recomb inant form of PRMT3 is inhibited by high concentrations of ZnCl2 as well as N-ethylmaleimide, reagents that can modify cysteine sulfhydryl groups. We found that we could distinguish PRMT family members by their sensitivity to these reagents; JBP1/PRMT5 and HsI7 methyltransferases were inhibited in a similar manner as PRMT3, whereas Rmt1, PRMT1, and CARM1/PRMT4 were not aff ected. We were also able to define differences in these enzymes by their se nsitivity to inhibition by Tris and free arginine. Finally, we found that t he treatment of RAT1 cell extracts with N-ethylmaleimide leads to a loss of the major PRMT1-associated activity that was immune to inhibition under th e same conditions as a GST fusion protein. These results suggest that nativ e forms of PRMTs can have different properties than their GST-catalytic cha in fusion protein counterparts, which may lack associated noncatalytic subu nits.