Human Thg1 displays tRNA-inducible GTPase activity

tRNA(His) guanylyltransferase (Thg1) catalyzes the 3 '-5 ' incorporation of guanosine into position -1 (G-1) of tRNA(His). G-1 is unique to tRNA(His) and is crucial for recognition by histidyl-tRNA synthetase (HisRS). Yeast Thg1 requires ATP for G-1 addition to tRNA(His) opposite A73, whereas archaeal Thg1 requires either ATP or GTP for G-1 addition to tRNA(His) opposite C73. Paradoxically, human Thg1 (HsThg1) can add G-1 to tRNAs(His) with A73 (cytoplasmic) and C73 (mitochondrial). As N73 is immediately followed by a CCA end (positions 74-76), how HsThg1 prevents successive 3 '-5 ' incorporation of G-1/G-2/G-3 into mitochondrial tRNA(His) (tRNA(m)(His)) through a template-dependent mechanism remains a puzzle. We showed herein that mature native human tRNA(m)(His) indeed contains only G-1. ATP was absolutely required for G-1 addition to tRNA(m)(His) by HsThg1. Although HsThg1 could incorporate more than one GTP into tRNA(m)(His)in vitro, a single-GTP incorporation prevailed when the relative GTP level was low. Surprisingly, HsThg1 possessed a tRNA-inducible GTPase activity, which could be inhibited by ATP. Similar activity was found in other high-eukaryotic dual-functional Thg1 enzymes, but not in yeast Thg1. This study suggests that HsThg1 may downregulate the level of GTP through its GTPase activity to prevent multiple-GTP incorporation into tRNA(m)(His).