{'fraction_non_alphanumeric': 0.03517877739331027, 'fraction_numerical': 0.01903114186851211, 'mean_word_length': 4.941780821917808, 'pattern_counts': {'":': 0, '<': 0, '<?xml version=': 0, '>': 0, 'https://': 0, 'lorem ipsum': 0, 'www.': 0, 'xml': 0}, 'pii_count': 0, 'substrings_counts': 0, 'word_list_counts': {'cursed_substrings.json': 0, 'profanity_word_list.json': 0, 'sexual_word_list.json': 0, 'zh_pornsignals.json': 0}}
Preferred Sequences within a Defined Cleavage Window Specify DNA 3′ End-directed Cleavages by Retroviral RNases H*

The RNase H activity of reverse transcriptase carries out three types of cleavage termed internal, RNA 5′ end-directed, and DNA 3′ end-directed. Given the strong association between the polymerase domain of reverse transcriptase and a DNA 3′ primer terminus, we asked whether the distance from the primer terminus is paramount for positioning DNA 3′ end-directed cleavages or whether preferred sequences and/or a cleavage window are important as they are for RNA 5′ end-directed cleavages. Using the reverse transcriptases of human immunodeficiency virus, type 1 (HIV-1) and Moloney murine leukemia virus (M-MuLV), we determined the effects of sequence, distance, and substrate end structure on DNA 3′ end-directed cleavages. Utilizing sequence-matched substrates, our analyses showed that DNA 3′ end-directed cleavages share the same sequence preferences as RNA 5′ end-directed cleavages, but the sites must fall in a narrow window between the 15th and 20th nucleotides from the recessed end for HIV-1 reverse transcriptase and between the 17th and 20th nucleotides for M-MuLV. Substrates with an RNA 5′ end recessed by 1 (HIV-1) or 2–3 (M-MuLV) bases on a longer DNA could accommodate both types of end-directed cleavage, but further recession of the RNA 5′ end excluded DNA 3′ end-directed cleavages. For HIV-1 RNase H, the inclusion of the cognate dNTP enhanced DNA 3′ end-directed cleavages at the 17th and 18th nucleotides. These data demonstrate that all three modes of retroviral RNase H cleavage share sequence determinants that may be useful in designing assays to identify inhibitors of retroviral RNases H.