Catalyzes conversion of folates to polyglutamate derivatives allowing concentration of folate compounds in the cell and the intracellular retention of these cofactors, which are important substrates for most of the folate-dependent enzymes that are involved in one-carbon transfer reactions involved in purine, pyrimidine and amino acid synthesis. Unsubstituted reduced folates are the preferred substrates. Metabolizes methotrexate (MTX) to polyglutamates. (6S)-5,6,7,8-tetrahydrofolyl-(gamma-L-Glu)(n) + ATP + L-glutamate = (6S)-5,6,7,8-tetrahydrofolyl-(gamma-L-Glu)(n+1) + ADP + H(+) + phosphate A monovalent cation. K(+) is most effective, followed by NH4(+) and Rb(+). Na(+), Li(+) and Cs(+) are ineffective. Activated by 10 mM sodium bicarbonate. Optimum pH is 9.6 (isoform 2). Cofactor biosynthesis; tetrahydrofolylpolyglutamate biosynthesis. Monomer. Produced by alternative initiation at Met-43 of isoform 1. Produced by alternative splicing of isoform 1. Produced by alternative splicing of isoform 1. Belongs to the folylpolyglutamate synthase family. Truncated N-terminus.