Gene, 123 (1993) 289-290

0 1993 Elsevier Science Publishers B.V. All rights reserved. 0378-l I19/93/$06.00

GENE 06870

289

Nucleotide sequence of the cDNA encoding mouse thyroid peroxidase

(Recombinant DNA; sequence homology; TPO activity; proximal His)

Tomio Kotani”, Kazumi Umeki”, Ikuo Yamamotoa, Masanori Takeuchi”, Shinji Takechib, Tatsuo Nakayamab and Sachiya Ohtaki”

Departments of”Laboratory Medicine and bBiochemistry, Miyazaki Medical College, Kiyotake, Miyazaki 889-16, Japan

Received by H. Yoshikawa: 24 August 1992; Revised/Accepted: 16 September/22 September 1992; Received at publishers: 2 October 1992

SUMMARY

The nucleotide (nt) sequence of the cDNA encoding mouse thyroid peroxidase (TPO) has been determined. The TPO cDNA is 3281 nt long and its open reading frame encodes a protein composed of 914 amino acids (aa), including a putative initiation Met. The mouse TPO cDNA shows 75.3, 70.8, and 93.5% homology in the coding nt sequence with human, porcine, and rat TPO cDNAs, respectively. In the aa sequence, mouse TPO shows 73.4, 68.4, and 93.9% homology with human, porcine, and rat TPOs, respectively. Specifically, the aa sequence surrounding the proximal His residue, probably essential for TPO activity, is well conserved among these four organisms.

TPO is a key enzyme in thyroid hormone synthesis. Recently, TPO has been identified not only as a major thyroid microsomal antigen in humoral immunity of autoimmune thyroid diseases (Czarnocka et al., 1985; Portmann et al., 1985; Kotani et al., 1986), but also as a thyroiditogenic antigen in experimental thyroiditis in- duced in C57BL/6 mice (Kotani et al., 1990). To study the role of TPO in mouse thyroid autoimmunity, we have cloned and sequenced the cDNA for mouse TPO from a C57BL/6 thyroid cDNA library using human TPO cDNA (Kimura et al., 1987) as a probe.

The mouse TPO cDNA is 3281 nt long (Fig. 1); this size is virtually equal to that of mouse TPO mRNA ob- tained by Northern blotting (data not shown). The cDNA consists of the 73-nt 5’-noncoding region, the 2742-nt

Correspondence to: Dr. S. Ohtaki, Department of Laboratory Medicine,

Miyazaki Medical College, Kiyotake, Miyazaki 889-16, Japan. Tel. (81-

985)85-3776; Fax @l-985)85-4709.

Abbreviations: aa, amino acid(s); cDNA, DNA complementary to

mRNA; nt, nucleotide(s); TPO, thyroid peroxidase; TPO, gene (DNA)

encoding TPO.

coding region encoding a 914-aa protein including a puta- tive Met start codon, and the 466-nt 3’-noncoding region. Within the 3’-untranslated region, it possesses a possible polyadenylation signal of the form ATTAAA located at positions 3259-3264 as well as poly(A) stretch. In coding nt sequence, the mouse TPO cDNA shows 75.3,70.8, and 93.5% homology with human (Kimura et al., 1987), por- cine (Magnusson et al., 1987) and rat (Derwahl et al., 1989) TPO cDNAs, respectively.

When the deduced aa sequences are compared, the mouse TPO shows 73.4, 68.4, and 93.9% homology with human, porcine, and rat TPOs, respectively. A His4” residue is reported to be essential for human TPO activity (Kimura and Ikeda-Saito, 1988). The His residue is con- served among three other species including the mouse (Fig. 2). Furthermore, the aa sequence around the proxi- mal His residue is also conserved compared with the re- maining aa sequences among the known four TPOs. These findings suggest that this particular His residue, proposed to be essential for the human TPO, has com- monly a key role for TPO activity.

290

CACAGAGAA~A~AC~~AG~GGTG~A~ATC~~G~TTC~T~CG~~G~~GAGAAAAGGA~ACGTTCAGC~CTAGA

ATGAGAACA~TTGGAGCTATGG~AATAATG~TGGTGGTTATGGGAACIG~AAT~TTCCTCT~TTTTAT~CTGAGAAGCAGAGACATCTTGTGTGGGAAGACCATGAAGTCCCATGTTATC MRTLGAMAIMLVVMGTVIFLSFILRSRDILCGKTMKSHVl

AGTGCTGTGGAAACGAGCCAGCTCATGGTGGACCATGCAGTCTACAACACCATGAAAAGAAACCTCAAGAAAAGGGAAGTCCTTTCTCCAGCCCAGCTTCTCTCTTTCTTTAAGCTGCCC SAVETSDLMVDHAVYNTMKRNLKKREVLSPAGLLSFFKLP

GAGTCCACCAGTGGGGCTATTTCCCGAGCAGCAGAGATTATGGAAACATCAATACAAGTCATGAAACGTGAACAGTCACAGTTCTCCACGGATGCCTTATCAGCAGACATTCTGGGCACA ESTSGAISRAAEIMETSIPVMKREOSOFSTDALSADILGT

ATTGCCAACCTGTCAGGATGCTTGCCTTTCATGCTGCCACCAAGATGTCCTGACACCTGCCTGGCAAATAAGTACCGGCCCATCACAGGGGCGTGCAACAATAGAGATCACCCCAGATGG IANLSGCLPFMLPPRCPDTCLANKYRPITGACNNRDHPRW

GGAGCCTCCAACACAGCCCTAGCAAGATGGCTGCCTCCTGTCTATGAAGATGGCTTCAGTCAGCCCAAAGGCTGGAACCCTAATTTCTTATACCACGGCTTCCCACTGCCCCCGGTACGG GASNTALARWLPPVYEDGFSGPKGWNPNFLYHGFPLPJ'VR

GAAGTGACAAGGCACCTCATTCAAGTTTCCAATGAGCCTGCGATCATGACATTGCTCTCACACCACAG EVTRHLIGVSNEAVTEDDPYSDFLPVWGQYIDHDIALTPII

AGCACTAGCACAGCAGCCTTCTGGGGAGGTGTCGACTGCCAGCTGACCTGTGAGAACCAAAATCCTTGCTTCCCCATACAGCTTCCCTCAAACTCCTCAGGGACCACTGCATGCCTGCCT STSTAAFWGGVDCGLTCENPNPCFPIGLPSNSSGTTACLP

TTCTACCGCTCCTCCGCCGCTTGTGGCACTGGGGACCAAGGTGCTCTCTTTGGCAACCTGTCTGCAGCCAATCCGAGGCAGCAGATGAATGGCTTGACCTCCTTCCTTGATGCTTCCACT FYRSSAACGTGDGGALFGNLSAANPRGGMNGLTSFLDAST

GTGTACGGCAGCTCCCCTGGCGTTGAGAAGCAGCTGCGCAACTGGAGCAGCTCGGCAGGACTGCTGCGTGTCAACACTCTCCACCTAGATGCTGGCCGTGCCTACCTGCCCTTCGCAACA VYGSSPGVEKGLRNWSSSAGLLRVNTLHLDAGRAYLPFAT

GCCGCCTGCGCTCCAGAGCCTGGTACCCCACGCACCAACCGCACGCCCTGCTTCCTGGCTGGAGACGGTCGCGCCAGTGAGGTCCCTGCCCTGGCAGCGGTACACACCTTGTGGCTGCGC AACAPEPGTPRTNRTPCFLAGDGRASEVPALAAVHTLWLR

GAGCACAACCGCCTGGCTTCGGCCTTCAAGGCCATTAACAAGCACTGGAGCGCCAACACTGCCTACCAGGAGGCGCGCAAGGTGGTAGGGGCACTGCACCAGATCATCACCATGAGGGAT EHNRLASAFKAINKHWSANTAYIIEARKVVGALHGIITMRD

TATATCCCCAAAATCCTGGGTCCTGATGCCTTCAGGCAGTATGTGGGCCCCTATGAAGGCTACAACCCCACTGTGAACCCTACTGTGTCCAACATCTTCTCCACTGCTGCCTTTCGCTTT YIPKILGPDAFRGYVGPYEGYNPTVNPTVSNIFSTAAFRF

GGCCATGCCACAGTCCATCCACTGGTGAGACGGCTAAACACTGACTTCCAGGAGCACACAGAGCTCCCCAGGTTGCAGCTGCGTGATGTCTTCTTCAGACCCTGGAGGCTTATCCAGGAA GHATVHPLVRRLNTDFGEHTELPRLGLRDVFFRPWRLIPE

GGTGGTTTGGATCCGATAGTGAGAGGCCTCCTGGCGAGAGCAGCCAAGCTGCAAGTCCAAGGGCAGCTGATGAATGAGGAGCTGACCGAGAGGCTCTTCGTGCTGTCTAACGTGGGCACC GGLDPIVRGLLARAAKLDVilGOLMNEELlERLFVLSNVGT

TTGGATCTGGCATCACTGAACTTGCAGAGGGGCCGGGATCATGGCTTACCAGACTACAATGAATGGAGAGAGTTCTGTGGCTTGTCACGCCTGGAGACACCAGCTGAGCTGAACAAGGCC LDLASLNLGRGRDHGLPDYNEWREFCGLSRLETPAELNKA

ATTGCCAACAGAAGCATGGTCAACAAGATAATGGACTTATACAAGCATGCTGACAACATTGACGTCTGGTTGGGAGGCTTGGCTGAAAAGTTCTTGCCGGGGGCCCGCACTGGTCCTCTG IANRSMVNKIMDLYKHADNIDVWLGGLAEKFLPGARTGPL

TTTGCATGTATCATTGGGAAGCAGATGAAGGCTCTGAGGGATGGGGACAGGTTTTGGTGGGAGAACACCAATGTCTTCACAGACGCTCAGAGGCAGG~CTAG~AAGCATTCACTACCT FACIIGKQMKALRDGDRFWWENTNVFTDAGRGELEKHSLP

CGGGTCATCTGTGACAACACTGGACTCACCAGAGTACCTGTGGATGCCTTCCGTATTGGAAAGTTCCCCCAAGACTTTGAATCCTGTGAGGACATCCCGAGCATGGACCTTGAACTATGG RVICDNTGLTRVPVDAFRIGKFPGDFESCEDIPSMDLELW

AGGGAGACCTTCCCACAAGATGACAAGTGTGTCTTCCCAGAGGAGGTGGACAATGGGAACTTTGTGCACTGTGAAGAGTCTGGGAAGCTGGTGCTTGTATACTCCTGTTTCCATGGATAC RETFPGDDKCVFPEEVDNGNFVHCEESGKLVLVYSCFHGY

AAGCTGCAGGGCCAGGAGCAGGTCACATGTACCCAGAAGGGATGGGACTCCGAACCTCCTGTCTGTAAAGATGTTAATGAGTGTGCAGATCTGACACACCCACCTTGCCACCCCTCTGCA KLGGGEGVTCTDKGWDSEPPVCKDVNECADLTHPPCHPSA

CAGTGCAAGAACACCAAGGGAAGCTTCCAGTGTGTGTGCACAGACCCCTATGTGCTAGGTGAGGATGAGAAGACCTGCATAGATTCTGGCAGGCTACCTCGGGCATCCTGGGTCTCCATT GCKNTKGSFGCVCTDPYVLGEDEKTCIDSGRLPRASWVSI

GCACTGGGTGCACTTCTCATTGGTGGTTTGGCCAGTCTCACCTGGATAGTAATTTGCAGGTGGACACATGCTGATAAGAAAGCCACATTGCCGAT~CAGAGAGAGTGACCACCCAGICA ALGALLIGGLASLTWIVICRWTHADKKATLPITERVTTGS

GGATGCAGAAAGAGTCAGGGGAGGGGGATTTCACCACACAAGGCCGCAGCTCAAGACACTGGACAGGAACCTGCAAGTGGATCCAGGGTCCTCCTGTGCGAATAGAGGTCCTCACTGCTT GCRKSQGRGISPHKAAAQDTGPEPASGSRVLLCE***

TGGAGCCAGACATTGGCAAAGGCAAGTCTCAAGCTGCCTGGGCAAAGACATGACACGTGTTGGAGTCAGAGACTTGAGGACACCAGATGCTGAGTCTTATCATCAGCCCAAGGCTGCAGT AGCTTCCATCTCATGTGTTTCCACAGGAGCAGTGCAGGCCAGACCGTGCTAATGCCTCTCCTACGCACTAAGGCTAAIGGATGCGTGGGAGGAAGGCTATGAGAATCACATMGCCIGAA AGTCCTGAGAGACGCTGGCATAGACTCCAGATCCTTCTAGACAAGTTCATGTTCATTCAGAATCAGCAAGGCTCTTTCAAATACACTCA TTCTCTCGTTACGGTCCAGTCACCACAAAG CACAGGCGAAGAAATGCTATAGCATTTTGCTGGTCTGGCTCAGATATCCTCCTCCTCTAGGCTTGC~GACATATTTACATATGA 10) 7

Fig. 1. The nt sequence of mouse TPO cDNA and the deduced aa sequence. Asterisks mark the stop codon. The possible polyadenylation signal is

underlined. EMBL act. No.: X60703.

House “’ PC PLA G DGRA SEV PALAAV HT LW LR EHN R LA 1o8 Human a88 P C P L A G D G R A S E V Pi’s:’ L ,:Ti A LI: H’T L W L R E H N R L A ‘I’ Porcine”’ P C P L A G D i S’ R A S E V R :;Gi: L P.: A ,I’ H T L W L R E H N R L A ‘On Rat “’ PC PLA GDG RA S E V PA LAAVHT LW LR EHN R LA aB

Fig. 2. Comparison of aa sequences around the proximal His residue. The aa are numbered for each of these TPOs and gaps have been introduced

to optimize alignment. Shaded boxes, aa different from those of mouse TPO. Asterisk marks the proximal His.

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