D-13-AAA; (B) R-13-GUUGU; (C) R-13-UCUCG; and (D) R-13-UGACA. In the control reactions, fluorescent intensities in the presence of 1 U of RNase I (blue) ...
Toxins 2016, 8, x
S1 of S1
Supplementary Materials: AAU‐Specific RNA Cleavage Mediated by MazF Toxin Endoribonuclease Conserved in Nitrosomonas europaea Tatsuki Miyamoto 1,2, Akiko Yokota 2, Satoshi Tsuneda 1,* and Naohiro Noda 1,2,*
Figure S1. MazFNE1181‐mediated sequence‐specific RNA cleavage. Two hundred nanograms of MazFNE1181 (green) was incubated with 20 pmol of fluorescent‐modified oligonucleotides; (A) D‐13‐AAA; (B) R‐13‐GUUGU; (C) R‐13‐UCUCG; and (D) R‐13‐UGACA. In the control reactions, fluorescent intensities in the presence of 1 U of RNase I (blue) and in the absence of enzymes (yellow) at each time point (left) and end point (right) were measured.
Toxins 2016, 8, x
S1 of S2
Figure S2. Neutralization of MazFNE1181‐mediated RNA cleavage. Ten picomoles of MazFNE1181 were pre‐incubated with 2 (vermillion), 10 (purple), or 50 (gray) pmol of MazENE1182, and mixed with 20 pmol of DR‐13‐AAU. In the control reactions, fluorescent intensities in the presence of 1 U of RNase I (blue) and MazFNE1181 (yellow) and in the absence of enzymes (green) were measured. (A) Fluorescent intensities at each time point; (B) Fluorescent intensities at end point. Table S1. Twenty‐five sequences with MazFNE1181 cleavage
RNA Type
1000‐1
1000‐2
1000‐3
Rank
Position
1 2 3 4 5 1 2 3 4 5 1 2
725 332 188 392 703 71 192 461 277 628 533 808
Relative Coverage Increase 4.93 2.53 2.11 1.78 1.68 5.12 2.49 1.79 1.75 1.68 2.19 1.89 1
Coverage
Sequence (5′ to 3′) a
25,940 14,909 5407 22,871 5667 1151 3703 6739 5066 16,116 1472 1061
CCCAAATAGAC CAGAAATCACT CCTAAATGGAC CGCCAATCTCT CGAGAATCATG ACCGAATCCCT ATTTAATGTTC CTTCAATTTGT TCGTAATGGTT GCGCAAAGGAC AATGAATATCG TCCCAATTCAA
Toxins 2016, 8, x
1000‐4
1000‐5
S2 of S2
3 4 5 1 2 3 4 5 1 2 3 4 5
218 849 726 98 480 663 335 798 36 715 432 716 362
1.39 1.39 1.36 52.27 2.88 2.21 2.06 1.81 27.44 9.34 1.81 1.61 1.58
1067 1294 1015 9200 10,727 7650 4102 3956 1427 8969 18,470 14,443 9653
TTGAAATCACC TTCGAATTTCG ATTCAATCTAC TCCCAATAGTT CCTGAATACAC CCTTAATAAGC GCCCAATACGT AGCTAATCGGA TCGGAATCTTT CATGAAATGAA CGCGAAAGGAT ATGAAATGAAC TACGAATGGGC
a: Underlined letters represent the base with significant coverage increase. Table S2. Genes consisting of mer operon.
Locus
Gene Symbol
Product Name
Length (bp)
NE0842 NE0841 NE0840 NE0839 NE0838 NE2575 NE0843
merT merP merC merA merD merE merR
mercuric transport protein mercury scavenger protein mercury transport protein mercuric reductase transcriptional regulator, MerR family mercury resistance protein transcriptional regulator, MerR family
351 276 423 1686 366 237 426
2
Number of AAU Triplets 0 0 2 9 2 0 4
