Plays a central role in 2-thiolation of mcm(5)S(2)U at tRNA wobble positions of cytosolic tRNA(Lys), tRNA(Glu) and tRNA(Gln). Acts by mediating the C-terminal thiocarboxylation of sulfur carrier URM1. Its N-terminus first activates URM1 as acyl-adenylate (-COAMP), then the persulfide sulfur on the catalytic cysteine is transferred to URM1 to form thiocarboxylation (-COSH) of its C-terminus. The reaction probably involves hydrogen sulfide that is generated from the persulfide intermediate and that acts as a nucleophile towards URM1. Subsequently, a transient disulfide bond is formed. Does not use thiosulfate as sulfur donor; NFS1 probably acting as a sulfur donor for thiocarboxylation reactions. Prior mcm(5) tRNA modification by the elongator complex is required for 2-thiolation. May also be involved in protein urmylation