ACE2 orthologues in non-mammalian vertebrates (Danio, Gallus, Fugu, Tetraodon and Xenopus).

"topo cloning was employed to clone fragments around 1 1 kb for sequencing 2 3 human and mouse ace2 sequences (protein [mouse ace2 ncbi accession no aah26801 ] cdna [human ncbi accession no bc039902 and mouse ncbi accession no bc026801 ] and genomic dna [supplementary data 1]) were retrieved from the ncbi database.; the full length ace2 sequences of fugu (803 amino acids see supplementary data 1) tetraodon (803 a a; supplementary data 1) and gallus (808 a a; supplementary data 1) were identified in silico from available databases.; the full-length danio ace2 sequence was then identified as an 807 a a (supplementary data 1) polypeptide.; we found that the polyadenylation signal was680 bp downstream of the stop codon and the frog ace2 encodes859 a a (supplementary data 1) residues with an extra 54 a a (compared to human ace2) located in the cytoplasmic domain.; table 1 exon-intron structures exons (coding sequences) introns (intronic regions) h a ace2 and m ace2 g ace2 x ace2 f ace2 and t ace2 h ace2 m ace2 g ace2 x ace2 f ace2 t ace2 1 186 183 189 195 5722 12203 2220 21076 86 86 2 159 b 159 159 159 2522 3297 1785 679 100 91 3 94 94 94 94 372 1257 571 1901 335 315 4 144 144 144 144 2256 2762 456 939 132 150 5 113 113 113 116 1485 1402 681 628 75 79 6 106 109 109 106 2180 1538 708 292 353 511 7 98 98 89 89 4084 3782 1428 583 102 88 8 170 170 170 173 2905 1609 867 531 611 601 9 227 227 227 227 2278 2558 604 1037 348 260 10 145 145 145 145 2200 1561 1604 1312 126 103 11 99 99 99 99 1043 1088 668 169 152 95 12 123 123 123 123 404 428 319 493 148 86 13 173 173 179 179 1270 1492 704 820 796 867 14 59 59 62 59 2468 2887 3951 4212 127 129 15 101 101 101 98 1356 2065 951 1385 88 95 16 117 117 111 117 2034 2925 1197 290 126 95 17 195 195 195 195 2010 1665 609 1210 151 113 18 106 115 268 91 tbp c 2415 2424 2577 2409 36589 44519 19323 37557 3856 3764 the number of nucleotides in each exon and intron are shown a ace2 s are prefixed by h (human) m (mouse) g (chicken) x ( xenopus ) f ( fugu ) and t ( tetraodon ) b high-lighted nucleotide numbers indicating the exon among all ace2 s are same c total base pairs3 2 we determined the exon-intron structure of the chicken frog and pufferfish ace2 genes by mapping the cdnas (supplementary data 1) to their respective genomic sequence.; the xenopus has the longest ace2 with 2477 bp (supplementary data 1) and the shortest 2409 bp are found in the pufferfish (supplementary data 1).; locus positions in the chromosome or scaffold are underlined 3 4 supplementary data 2 presents the alignment of ace2s from human mouse chicken frog and teleosts ( fugu tetraodon and zebrafish) ace2 is a zinc metallo-carboxymonopeptidase and it is distinct from ace as it carries only a single catalytic domain to perform the monopeptidase activity.; a movie of a beating fluorescent heart from a transgenic frog (using human 0 6 kb putative ace2 promoter to drive the expression of egfp) is presented as supplementary data 3.; ten out of eighteen ace2 exons show length variations in the six animals however the exon boundaries in all six genes are identical as indicated by arrows in supplementary data 2.; we found seven cysteine residues conserved in all ace2s (supplementary data 2) suggesting that they may be responsible for maintaining the conformation stability ( ) or biological activities ( ) ace2 residues arg 273 (critical for substrate binding) his345 and his505 involved in catalysis as proven by site-directed mutagenesis ( ) are conserved in all 7 genes.; therefore since chicken ace2 contains a glu at position 31 (supplementary data 2) we hypothesize that this may be an obstacle in the binding between sars-cov s protein and chicken ace2.; appendix a supplementary data 1 online supplementary data 1 ace2 orthologues in non-mammalian vertebrates (danio gallus fugu tetraodon and xenopus) online supplementary data 1 ace2 orthologues in non-mammalian vertebrates (danio gallus fugu tetraodon and xenopus)."

"We thank the IMCB DNA Sequencing Facility for its help in sequencing. This project was supported by the Agency for Science and Technology (A⁎STAR) Singapore."