JPH03155795A - Mouse-interleukin-6 receptor protein - Google Patents
Mouse-interleukin-6 receptor proteinInfo
- Publication number
- JPH03155795A JPH03155795A JP1292230A JP29223089A JPH03155795A JP H03155795 A JPH03155795 A JP H03155795A JP 1292230 A JP1292230 A JP 1292230A JP 29223089 A JP29223089 A JP 29223089A JP H03155795 A JPH03155795 A JP H03155795A
- Authority
- JP
- Japan
- Prior art keywords
- mouse
- receptor
- dna
- amino acid
- receptor protein
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
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Abstract
Description
【発明の詳細な説明】
〔産業上の利用分野〕
本発明はマウスIL−6レセプター蛋白質および該蛋白
質をコードするDNA、さらには該蛋白質を遺伝子工学
的に生産するための手段および方法に関するものである
。Detailed Description of the Invention [Field of Industrial Application] The present invention relates to mouse IL-6 receptor protein, DNA encoding the protein, and means and methods for producing the protein through genetic engineering. be.
IL−6は、免疫、造血、炎症という生体防御系におい
て重要な役割を果たしていることを特徴とする、生体の
増殖分化に広く関与するタンパク質である。一方、IL
−6の異常産生が種々の自己免疫疾患の病因因子である
可能性が報告されている(厚木、平野、Ann、Rev
、Immunol、、 5. p435゜1988年参
照)。IL-6 is a protein widely involved in the proliferation and differentiation of living organisms, which is characterized by playing an important role in the biological defense systems of immunity, hematopoiesis, and inflammation. On the other hand, IL
It has been reported that abnormal production of -6 may be an etiological factor for various autoimmune diseases (Atsugi, Hirano, Ann, Rev.
, Immunol, 5. (See p. 435゜1988).
IL−6のシグナル伝達経路は、以下のように解明され
てきた。まずIL−6と特異的に結合する細胞膜上のI
L−6レセプターが、田賀らにより解析され、各細胞上
の数、IL−6との結合定数が報告された(J、Exp
、Med、、 196. p967、1987年参照)
。次にヒ)IL−6レセプターのcDNAが出端らによ
り単離され、1次構造が決定された(Science、
241. p825.1988年参照)。さらにIL
−6のシグナル伝達に関与する細胞膜上の別の蛋白質が
田賀らにより発見された(Cell、 58゜p573
.1989年参照)。The IL-6 signal transduction pathway has been elucidated as follows. First, I on the cell membrane that specifically binds to IL-6.
The L-6 receptor was analyzed by Taga et al., and the number on each cell and the binding constant with IL-6 were reported (J, Exp
,Med,, 196. (see p.967, 1987)
. Next, the cDNA of the IL-6 receptor was isolated by Ida et al., and the primary structure was determined (Science,
241. p825.1988). Further IL
Another protein on the cell membrane involved in the -6 signal transduction was discovered by Taga et al. (Cell, 58゜p573
.. (see 1989).
生体内で多様な生理活性を強く発揮するIL−6の作用
を人為的に調節することは、各種疾患の新しい治療のメ
カニズムとして期待されている。Artificial regulation of the effects of IL-6, which strongly exerts various physiological activities in vivo, is expected to be a new therapeutic mechanism for various diseases.
例えば、IL−6の骨髄細胞増殖効果や血小板増加効果
は放射線等の癌治療の効果を高める。一方、各種自己免
疫疾患では、その病因と考えられるIL−6作用の抑制
が、症状の軽減につながる。For example, the bone marrow cell proliferation effect and platelet increase effect of IL-6 enhance the effects of cancer treatments such as radiation. On the other hand, in various autoimmune diseases, suppression of IL-6 action, which is thought to be the cause of the disease, leads to alleviation of symptoms.
前者の目的には、遺伝子工学的に作成されたヒトIL−
6が、後者の目的には、遺伝子工学的に作成された細胞
表層より離脱したく可溶性)ヒトIL−6レセプターや
ヒトIL−6に対する中和抗体が考えられるが、さらに
他の合成物質や天然物質にもIL−6の作用を調節する
物質の存在が期待される。For the former purpose, genetically engineered human IL-
6. For the latter purpose, genetically engineered human IL-6 receptors (which are soluble and want to be released from the cell surface) and neutralizing antibodies against human IL-6 can be considered, but other synthetic substances and natural The existence of substances that regulate the effects of IL-6 is also expected.
可溶性ヒトIL−6レセプターをIL−6作用を調節す
る薬剤として開発するためには、マウス等の実験動物に
投与して、その効果を調べることが必須となる。しかし
、ヒト可溶性IL−6レセプターをマウス等の実験動物
に投与しても、実験動物由来のIL−6と特異的に結合
することが期待できない。さらには、ヒト可溶性IL−
6レセプターの実験動物に対する抗原性も問題となる。In order to develop soluble human IL-6 receptor as a drug that modulates IL-6 action, it is essential to administer it to experimental animals such as mice and examine its effects. However, even if human soluble IL-6 receptor is administered to experimental animals such as mice, it cannot be expected to specifically bind to IL-6 derived from the experimental animal. Furthermore, human soluble IL-
The antigenicity of the 6 receptor to experimental animals is also a problem.
実験動物の中では、マウスが、取り扱いの容易さにおい
て、さらにはIL−5過剰産生により自己免疫疾患にな
るストレインが確立されているという点において、最も
その使用が望まれる。しかし、マウス可溶性IL−6レ
セプターを遺伝子工学的に生産するには、マウスIL−
6レセプターをコードするDNA配列が必須である。Among experimental animals, the use of mice is most desirable because they are easy to handle and furthermore, there is a well-established strain of autoimmune disease due to overproduction of IL-5. However, to produce mouse soluble IL-6 receptor by genetic engineering, mouse IL-6 receptor
A DNA sequence encoding the 6 receptor is essential.
従って、本発明はマウスIL−6レセプター及びそれを
コードするDNA並びに該DNAを用いる該レセプター
の製造方法を提供しようとするものである。Therefore, the present invention provides a mouse IL-6 receptor, a DNA encoding it, and a method for producing the receptor using the DNA.
〔課題を解決するための手段〕
上記の目的を達成するために、本発明者は、IL−6レ
セプターについて鋭意研究を行った結果、マウスIL−
6レセプターをコードするDNA配列を明らかにし、こ
の知見をもとに遺伝子工学的にIL−6レセプターを生
産する手段および方法を完成した。[Means for Solving the Problems] In order to achieve the above object, the present inventor conducted intensive research on the IL-6 receptor, and as a result, the mouse IL-6 receptor was discovered.
The DNA sequence encoding the IL-6 receptor was revealed, and based on this knowledge, a means and method for producing the IL-6 receptor using genetic engineering was completed.
すなわち本発明は、マウスIL−6と特異的に結合する
マウスIL−6レセプター;マウスIL−6レセプター
をコードするDNA配列;組換微生物または培養細胞中
で前記DNA配列を発現し得る複製可能な発現ベクター
;前記発現ベクターにより形質転換された微生物または
培養細胞;および前記微生物または培養細胞においてマ
ウスIL−6レセプターをコードするDNA配列を発現
させることを特徴とするマウスIL−6レセプターの生
産方法、を提供するもであり、以下詳細1ご言桑明する
。Specifically, the present invention provides a mouse IL-6 receptor that specifically binds mouse IL-6; a DNA sequence encoding the mouse IL-6 receptor; a replicable microorganism capable of expressing said DNA sequence in a recombinant microorganism or cultured cells. an expression vector; a microorganism or cultured cell transformed with the expression vector; and a method for producing a mouse IL-6 receptor, which comprises expressing a DNA sequence encoding the mouse IL-6 receptor in the microorganism or cultured cell. We will provide you with the details below.
1、 マウスIL−6レセプター
マウスIL−6レセプターは、マウス由来の細胞膜上に
存在し、マウスIL−6と特異的に結合するタンパク質
である。詳しくは次の式(I)Met Leu Thr
Val Gly Cys Thr Leu Leu
VatAla Leu Leu Ala Al
a Pro Ala Val Ala Le
uVal Leu Gly Ser Cys Arg
Ala Leu Glu ValAla Asn Gl
y Thr Val Thr Ser Leu Pro
Gly^1a Thr Val Thr L
eu Ile Cys Pro Gly L
ysGlu Ala Ala Gly Asn Vat
Thr [1e His TrpVal Tyr S
er Gly Ser Gln Asn Arg Gl
u TrpThr Thr Thr Gly Asn
Thr Leu Val Leu ArgAsp Va
l Gfn Leu Ser Asp Thr Gly
Asp TyrLeu Cys Ser Leu A
sn Asp )lis Leu Vat GlyTh
r Val Pro Leu LeuGlu Glu
Pro Lys Leu^sn Pro Leu
Val AsnArg Pro Ser Ser
ThrLys Ala Val Leu PheTh
r Thr Asn Gly LysPro Cys
Gln Tyr 5erPhe Ser Cys Gi
n ValAsp Lys Vat Tyr HtsV
al Ala Asn Ser ValHis Asn
Glu Ala PheVal Gln Pro A
sp Pr。1. Mouse IL-6 Receptor Mouse IL-6 receptor is a protein that exists on mouse cell membranes and specifically binds to mouse IL-6. For details, use the following formula (I) Met Leu Thr
Val Gly Cys Thr Leu Leu
VatAlaLeuLeuAlaAl
a Pro Ala Val Ala Le
uVal Leu Gly Ser Cys Arg
Ala Leu Glu ValAla Asn Gl
y Thr Val Thr Ser Leu Pro
Gly^1a Thr Val Thr L
eu Ile Cys Pro Gly L
ysGlu Ala Ala Gly Asn Vat
Thr [1e His TrpVal Tyr S
er Gly Ser Gln Asn Arg Gl
u TrpThr Thr Thr Thr Gly Asn
Thr Leu Val Leu ArgAsp Va
l Gfn Leu Ser Asp Thr Gly
Asp TyrLeu Cys Ser Leu A
sn Asp )lis Leu Vat GlyTh
r Val Pro Leu LeuGlu Glu
Pro Lys Leu^sn Pro Leu
Val AsnArg Pro Ser Ser
ThrLys Ala Val Leu PheTh
r Thr Asn Gly LysPro Cys
Gln Tyr 5erPhe Ser Cys Gi
n ValAsp Lys Vat Tyr HtsV
al Ala Asn Ser ValHis Asn
Glu Ala PheVal Gln Pro A
sp Pr.
Val Ser Ala Ile Pr。Val Ser Ala Ile Pr.
Leu Lys Vat Ser TrpTrp As
p Pro Ser TyrGin Leu Arg
Tyr ArgGlu Phe Thr Val L
euGln Tyr Gin Cys ValArg
Gly Val Lys HlsGly Lys
Glu Glu LeuVal Asp Vat
Pro Pr。Leu Lys Vat Ser TrpTrp As
p Pro Ser TyrGin Leu Arg
Tyr ArgGlu Phe Thr Val L
euGln Tyr Gin Cys ValArg
Gly Val Lys HlsGly Lys
Glu Glu LeuVal Asp Vat
Pro Pr.
Ser Cys Phe Arg Lys^1a li
e Cys Glu TrpPro Ser Pro
Thr ThrAla Lys Lys lie
Asn5er Asp Phe Gin VatG
in Gln Leu Lys 5erGlu lie
Leu Glu Glylie Val Ser L
eu CysGly Ser Lys Ser 5er
His Ser Leu Lys MetPro Al
a Asn Leu ValGly Arg Pro
Arg TrpGln His Pro Glu Th
rTyr Leu Leu Gin PhePro
Val Trp Ser LysLeu Leu
Pro Val Alalle tlis Asp
Ala LeuVat Val Gin Val A
rgAsp Leu Gly Gln TrpSer
Glu Trp Ser Pro Glu Vat T
hr Gly ThrPro Trp Ile Ala
Glu Pro Arg Thr Thr Pr。Ser Cys Phe Arg Lys^1a li
e Cys Glu TrpPro Ser Pro
Thr Thr Thr Ala Lys Lys lie
Asn5er Asp Phe Gin VatG
in Gln Leu Lys 5erGlu lie
Leu Glu Glylie Val Ser L
eu CysGly Ser Lys Ser 5er
His Ser Leu Lys MetPro Al
a Asn Leu ValGly Arg Pro
Arg TrpGln His Pro Glu Th
rTyr Leu Leu Gin PhePro
Val Trp Ser LysLeu Leu
Pro Val Allalle tlis Asp
Ala LeuVat Val Gin Val A
rgAsp Leu Gly Gln TrpSer
Glu Trp Ser Pro Glu Vat T
hr Gly ThrPro Trp Ile Ala
Glu Pro Arg Thr Thr Pr.
Ala Gly Ile Leu Trp Asn P
ro Thr Gin ValSer Vat Glu
Asp Ser Ala Asn 1lis Glu
AspGin Tyr Glu Ser Ser T
hr Glu Ala Thr 5erVal Le
u Ala Pro Vat Gln Glu Se
r Ser SerMet Ser Leu Pro
Thr Phe Leu Vat Ala GlyGl
y Ser Leu Ala Phe Gly Leu
Leu Leu CysVal Phe Ile I
le Leu Arg Leu Lys Gln Ly
sTrp Lys Ser Glu Ala Glu
Lys Glu Ser LysThr Thr Se
r Pro Pro Pro Pro Pro Tyr
5erLeu Gly Pro Leu L
ys Pro Thr Phe Leu L
euVal Pro Leu Leu Thr Pro
)!+s Ser Ser GlySer Asp
Asn Thr Val Asn His Ser C
ys LeuGly Val Ala Asp Ala
Gin Ser Pro Tyr AspAsn S
er Asn Arg Asp Tyr Leu Ph
e Pro Arg(C末端)
はグルタミン、Gluはグルタミン酸、Glyはグリシ
ン、Hisはヒスチジン、Ileはアスパラギン酸、C
ysはロイシン、Lysはリジン、Metはメチオニン
、Pheはフェニルアラニン、Proはヒスチジン、I
leはセリン、Thrはトレオニン、Trp はリジン
、Netはチロシン、そしてValはフェニルアラニン
、Proはプロリン、Serはセリン、Thr はトレ
オニン、Trpはトリプトファン、Tyrはチロシン、
そしてVal はバリン残基を示す。)で表されるアミ
ノ酸配列を有するものである。本発明では前記アミノ酸
配列を有する蛋白質の他、前記アミノ酸配列中のIL−
6との特異的な結合に寄与する部分を含有する蛋白質ま
たはポリペプチドであれば良い。すなわち前記アミノ酸
配列中の1個または複数個のアミノ酸残基が他のアミノ
酸残基により置換されているかまたは欠失もしくは付加
されているアミノ酸配列を有し、かつマウスIL−6と
特異的に結合する能力を保持している蛋白質またはポリ
ペプチドはすべて本発明のマウスIL−6レセプターの
範囲に含まれる。例えば、このような蛋白質として、前
記アミノ酸配列中のtL−6との結合に寄与するアミノ
酸配列および/l、たはアミノ酸残基部分以外のアミノ
酸配列および/またはアミノ酸残基が置換、欠損、挿入
等により変化したもの、さらには前記アミノ酸配列のN
末端側および/またはC末端側にアミノ酸配列および/
またはアミノ酸残基が追加された蛋白質、あるいは例え
ばヒト成長ホルモン等の他の蛋白質等が融合したもので
あっても良い。Ala Gly Ile Leu Trp Asn P
ro Thr Gin Val Ser Vat Glu
Asp Ser Ala Asn 1lis Glu
AspGin Tyr Glu Ser Ser T
hr Glu Ala Thr 5erVal Le
u Ala Pro Vat Gln Glu Se
r Ser Ser Met Ser Leu Pro
Thr Phe Leu Vat Ala GlyGl
y Ser Leu Ala Phe Gly Leu
Leu Leu CysVal Phe Ile I
Le Leu Arg Leu Lys Gln Ly
sTrp Lys Ser Glu Ala Glu
Lys Glu Ser LysThr Thr Se
r Pro Pro Pro Pro Pro Tyr
5erLeu Gly Pro Leu L
ys Pro Thr Phe Leu L
euVal Pro Leu Leu Thr Pro
)! +s Ser Ser GlySer Asp
Asn Thr Val Asn His Ser C
ys LeuGly Val Ala Asp Ala
Gin Ser Pro Tyr AspAsn S
er Asn Arg Asp Tyr Leu Ph
e Pro Arg (C-terminus) is glutamine, Glu is glutamic acid, Gly is glycine, His is histidine, Ile is aspartic acid, C
ys is leucine, Lys is lysine, Met is methionine, Phe is phenylalanine, Pro is histidine, I
le is serine, Thr is threonine, Trp is lysine, Net is tyrosine, Val is phenylalanine, Pro is proline, Ser is serine, Thr is threonine, Trp is tryptophan, Tyr is tyrosine,
And Val indicates a valine residue. ) has the amino acid sequence represented by In the present invention, in addition to proteins having the above amino acid sequence, IL-
Any protein or polypeptide containing a portion that contributes to specific binding to 6 may be used. That is, the amino acid sequence has an amino acid sequence in which one or more amino acid residues are substituted with other amino acid residues, deleted or added, and specifically binds to mouse IL-6. All proteins or polypeptides that retain the ability to do so are within the scope of the murine IL-6 receptor of the present invention. For example, in such a protein, the amino acid sequence and /l that contribute to binding to tL-6 in the amino acid sequence, or the amino acid sequence and/or amino acid residue other than the amino acid residue portion, are substituted, deleted, or inserted. etc., and furthermore, the N of the above amino acid sequence.
Amino acid sequence and/or on the terminal side and/or C-terminal side
Alternatively, it may be a protein with added amino acid residues, or a fusion of other proteins such as human growth hormone.
式(1)で示されるアミノ酸配列は460アミノ酸残基
からなり、N末端側から第2番目に位置するロイシンか
ら第19番目に位置するアラニンにかけてと、第358
番目に位置するセリンから第385番目に位置するロイ
シンにかけての部分に疎水性アミノ酸残基が位置してい
る、この2つの疎水性領域は、前者がシグナルペプチド
領域、後者が膜貫通領域であると考えられる。The amino acid sequence represented by formula (1) consists of 460 amino acid residues, from leucine located at the second position from the N-terminal side to alanine located at the 19th position, and from the 358th amino acid residue to the 19th position alanine.
Hydrophobic amino acid residues are located between serine at position 385 and leucine at position 385. These two hydrophobic regions are thought to be a signal peptide region and a transmembrane region, respectively. Conceivable.
2、 DNA配列
生体内で産生されるマウスIL−6レセプターをコード
するDNA配列は、詳しくは次の式(n)により示され
る。2. DNA Sequence The DNA sequence encoding the mouse IL-6 receptor produced in vivo is specifically represented by the following formula (n).
(5−)
ATCCTG ACC
GCCCTG CTG
GTCCTCGGG
GCA AAT GGC
GCCACCGTT
6^^GCA GCA
GTG TACTCT
AcT AcCACA
GACGTG CAG
TTA TGCTCC
ACT GTG CCC
GAG GAG CCC
AACCCCCTT
CGT CCG AGC
AAG GCT GTG
ACCACCAAC
CCCTGCCAG
TTCTCCTGC
GACAAA GTA
GTC’ GGCTGCACG
GCCGCG CCCGCG
AGCTGCCGCGCG
ACA GTG AC^^GC
ACCCTG ATT TGC
GGCAAT GTT ACC
GGCTCA CA^^AC
GGA AACACA CTG
CTCAGCGACACT
CTG AAT GAT CAC
TTG CTG GTG GAT
AAG CTCTCCTGC
GTCAACGCCATC
AGCACCCCCTCT
CTG TT丁 GCA AAG
GGG AAG AGT GAC
TAT TCT CAG CAGCAG GTG
GAG ATC
TACCACATA GTG
CTG TTG GTC
GTCGCG CTG
CTG GAG GTG
CTG CCA GGG
CCCGGG AAG
ATT CACTGG
AGA GAA TGG
GTT CTG AGG
GGG GACTAT
CTG GTG GGG
GTT CCCCCA
TTCCGG 八^G
TGT GAG TGG
CCA ACCACG
AAA ATCAAC
TTCCAG GTG
CTG AAA AGE:
CTG GAG GGT
TCA CTG TGC
GTT GCA AACAGT GTGCACAACG
AA GCG TTT
GTG CAG CCG GAT CCAGTA TC
AGCCATA CCT
CTCAAA GTCAGCTGG
TGG GACCCG AGT TACCAG CTT
CGA TACCGAG八G へTTCACG G
TG TTGCAG TACCAA TGCGTC
CGA GGA GTG AAG CACGGG AA
G GAG GAG CTTAGT GAA TGG
TCCCCACCT TGG 八TA GCA
G^6GCA GGA ATCCTCTGG
TCT GTT GAA GACTCTCAG TAC
GAA AGT TCTGTCCTCGCCCCA G
TG
AGG TCC(1:TG CCCACAGGA
AGCTTG GCG TTTGTCTTCATCAT
CCTG
GGA AGCAAG
CACAGCTTA
CCT GCCAAC
GGA AGG CCG
CAG CACCCT
TACTTG CTG
CCT GTA TGG
CTG CTCCCG
ATCCAT GAT
GTG GTCCAG
GACCTT GGC
GAG GTCACG
CCCAGG ACC
AACCCCACA
GCCAACCAC
ACA GAA GCA
CAA GAA TCC
TTCCTG GTA
GGG TTG CTT
AGA CTCAAG
TCCAGC
AAA ATG
CTT GTG
CGCTGG
GAG ACC
CAG TTC
TCA AAG
GTG GCC
GCCTTG
GTCCGT
CAG TGG
GGCACT
ACCCCG
CAG GTC
GAG GAT
ACG AGT
TCG TCC
GCT GGA
CTCTGT
CAG AAA
TGG AAG TCA GAG GCT GAG A
AG GAA AGCAAGACG ACCTCT C
CT CCA CCCCCA CCG TAT TCC
TTG GGCCCA CTG AAG CCG AC
CTTCCTT CTGGTT CCT CTCCTC
ACCCCA CACAGCTCT GGGTCT G
ACAAT ACCGTA AACCACAGCTGC
CTGGGT GTCAGG GACGCA CAG
AGCCCT TAT GACAACAGCAACAG
A GACTACTTA TTCCCCAGA(3−)
で表されるDNA配列を有するものである。本発明のD
NA配列は、式(n)のDNA配列の他、このDNA配
列中の1個または複数個のヌクレオシドが他のヌクレオ
シドにより置換されているかまたは欠失もしくは付加さ
れているDNA配列を有し、かつマウスIL−6と特異
的に結合する能力を有する蛋白質をコードしているDN
A配列をも含有する。例えば、前記lにおいて記載した
種々のアミノ酸配列を有する蛋白質をコードするDNA
配列があげられる。(5-) ATCCTG ACC GCCCTG CTG GTCCTCGGG GCA AAT GGC GCCACCGTT 6^^GCA GCA GTG TACTCT AcT AcCACA GACGTG CAG TTA TGCTCC ACT GTG CCC GAG GAG CCC AACCCCCT CGT CCG AGC AAG GCT GTG ACCACCAAC CCCTGCCAG TTCTCCTG GACAAA GTA GTC' GGCTGCACG GCCGCG CCCGCG AGCTGCCGCGCG ACA GTG AC^^GC ACCCTG ATT TGC GGCAAT GTT ACC GGCTCA CA^^AC GGA AACACA CTG CTCAGCGACACT CTG AAT GAT CAC TTG CTG GT G GAT AAG CTCTCCTGC GTCAACGCCATC AGCACCCCCTCT CTG TT Ding GCA AAG GGG AAG AGT GAC TAT TCT CAG CAGCAG GTG
GAG ATC TACCACATA GTG CTG TTG GTC GTCGCG CTG CTG GAG GTG CTG CCA GGG CCCGGG AAG ATT CACTGG AGA GAA TGG GTT CTG AGG GGG GA CTAT CTG GTG GGG GTT CCCCCA TTCCGG 8^G TGT GAG TGG CCA ACCACG AAA ATCAAC TTCCAG GTG CTG AAA AGE: CTG GAG GGT TCA CTG TGC GTT GCA AACAGT GTGCACAACG
AA GCG TTT GTG CAG CCG GAT CCAGTA TC
AGCCATA CCT CTCAA GTCAGCTG TGG GACCCG AGT TACCAG CTT
CGA TACCGAG 8G to TTCACG G
TG TTGCAG TACCAA TGCGTC CGA GGA GTG AAG CACGGG AA
G GAG GAG CTTAGT GAA TGG
TCCCCACCT TGG 8TA GCA
G^6GCA GGA ATCCTCTG TCT GTT GAA GACTCTCAG TAC
GAA AGT TCTGTCCTCGCCCCA G
TG AGG TCC (1:TG CCCACAGGA
AGCTTG GCG TTTGTCTTCATCAT
CCTG GGA AGCAAG CACAGCTTA CCT GCCAAC GGA AGG CCG CAG CACCCT TACTTG CTG CCT GTA TGG CTG CTCCCG ATCCAT GAT GTG GTCCAG GACCT T GGC GAG GTCACG CCCAGG ACC AACCCCACA GCCAAACCAC ACA GAA GCA CAA GAA TCC TTCCTG GTA GGG TTG CTT AGA CTCAAG TCCAGC AAA ATG CT T GTG CGCTGG GAG ACC CAG TTC TCA AAG GTG GCC GCCTTG GTCCGT CAG TGG GGCACT ACCCCG CAG GTC GAG GAT ACG AGT TCG TCC GCT GGA CTCTGT CAG AAA TG G AAG TCA GAG GCT GAG A
AG GAA AGCAAGACG ACCTCT C
CT CCA CCCCCA CCG TAT TCC
TTG GGCCCA CTG AAG CCG AC
CTTCCTT CTGGTT CCT CTCCTC
ACCCCA CACAGCTCT GGGTCT G
ACAAT ACCGTA AACCACAGCTGC
CTGGGT GTCAGG GACGCA CAG
AGCCCT TAT GACAACAGCAACAG
It has a DNA sequence represented by A GACTACTTA TTCCCCAGA (3-). D of the present invention
In addition to the DNA sequence of formula (n), the NA sequence has a DNA sequence in which one or more nucleosides in this DNA sequence are substituted with other nucleosides, or deleted or added, and DN encoding a protein that has the ability to specifically bind to mouse IL-6
It also contains the A sequence. For example, DNA encoding proteins having the various amino acid sequences described in section 1 above.
Arrays can be given.
これらのDNA配列は、例えはBALB/c等のマウス
の例えば膵臓細胞から、公知な方法で抽出した種々のメ
ツセンジャーRNAより、本発明により提供される式(
n)DNA配列をもとに作製したプローブ等を用いて目
的とするメツセンジャーRNAを抽出し、これをもとに
作製しても良いし、また例えば一部あるいは全部を本発
明をもとに化学的に合成しても良い。These DNA sequences are derived from the formula (
n) The desired metsenger RNA may be extracted using a probe etc. prepared based on the DNA sequence and prepared based on this, or for example, a part or all may be based on the present invention. It may also be chemically synthesized.
3、発現ベクター
本発明で提供されるIL−6レセプターをコードするD
NA配列を発現、すなわち生産し得る複製可能な発現ベ
クターは、前記2で説明されたようなりNA配列を発現
させ得るDNA配列と読取り可能に結合しているマウス
IL−6レセプターをコードするDNA配列および宿主
中でベクターDNAを複製するための複製オリジン等を
有し、選定した宿主を形質転換できるものであれば制限
なく適宜選定して使用できる。例えば宿主として大腸菌
等の細菌株を用いる場合は、pBR322,pBR32
7等のプラスミドが例示できる。また例えば、宿主とし
て哺乳動物等の培養細胞を用いる場合には、SV40ウ
ィルス由来のDNA複製オリジンを有するベクターが例
示できる。3. Expression vector D encoding the IL-6 receptor provided by the present invention
A replicable expression vector capable of expressing or producing an NA sequence comprises a DNA sequence encoding the mouse IL-6 receptor readably linked to a DNA sequence capable of expressing the NA sequence as described in 2 above. It can be appropriately selected and used without any restriction as long as it has a replication origin for replicating the vector DNA in the host and can transform the selected host. For example, when using a bacterial strain such as E. coli as a host, pBR322, pBR32
An example is a plasmid such as No. 7. For example, when cultured cells of a mammal or the like are used as a host, a vector having a DNA replication origin derived from the SV40 virus can be exemplified.
DNA配列を発現させるためのDNA配列の中でも、プ
ロモーター系は重要であり、しかも選定した宿主との関
係において適宜選定する必要である。宿主として大腸菌
を使用する場合のプロモーター系としては、乳糖プロモ
ーター系、トリプトファンプロモーター系、さらにはこ
れらのハイブリッドプロモーター系が例示できる。宿主
として哺乳動物由来の培養細胞を使用する場合のプロモ
ーター系としては、SV40プロモーター系、アデノウ
ィルスプロモーター系が例示できる。Among the DNA sequences for expressing a DNA sequence, the promoter system is important and needs to be selected appropriately in relation to the selected host. Examples of promoter systems when using Escherichia coli as a host include a lactose promoter system, a tryptophan promoter system, and a hybrid promoter system thereof. Examples of promoter systems when using mammalian-derived cultured cells as hosts include the SV40 promoter system and the adenovirus promoter system.
4、宿主
本発明では、特別の制限なしに通常の遺伝子工学的に蛋
白質を生産するために用いられる微生物または培養細胞
が使用できる。微生物としては、K−12・W−311
0等の大腸菌類、枯草菌類、酵母等を例示できる。培養
細胞としては、CO8細胞(猿の腎臓繊維芽細胞)・、
CHO細抱(チャイニーズハムスターの卵母細胞)等が
例示できる。4. Host In the present invention, microorganisms or cultured cells commonly used for producing proteins by genetic engineering can be used without any particular limitations. As microorganisms, K-12 and W-311
Examples include Escherichia coli, Bacillus subtilis, and yeast. Cultured cells include CO8 cells (monkey kidney fibroblasts),
An example is CHO cells (Chinese hamster oocytes).
5、形質転換された宿主を用いたマウスIL−6レセプ
ターの生産
前記3で説明したベクターにより形質転換された前記4
で説明された様な宿主を培養し、ベクターDNA中のマ
ウスIL−6レセプターをコードするDNA配列を発現
させることでマウスIL−6レセプターを生産すること
ができる。これはマウスIL−6レセプターをコードす
るDNA配列を結合した該DNA配列を発現させ得るD
NA配列中のプロモーター系の活性化により実現される
。5. Production of mouse IL-6 receptor using a transformed host
A mouse IL-6 receptor can be produced by culturing a host as described in 1. and expressing the DNA sequence encoding the mouse IL-6 receptor in the vector DNA. This is a DNA sequence capable of expressing a DNA sequence encoding the mouse IL-6 receptor.
This is achieved by activation of a promoter system in the NA sequence.
以下本発明をさらに詳細に説明するために実施例を示す
が、本発明はこれら実施例に限定されるものではない。Examples will be shown below to explain the present invention in more detail, but the present invention is not limited to these Examples.
実施例1. マウスIL−6レセプターcDNへの単離
一連の遺伝子組み換え操作方法(m−RNAの抽出、D
NAの制限酵素による切断等)は、マニアナイスらの方
法(Molecular Cloning、 Co1d
SprrngHarbor Laboratory
1982年参照)、バーウィンらの方法(DNAclo
ning、 a Practical Approac
h vol。Example 1. Isolation of mouse IL-6 receptor cDNA A series of genetic recombination procedures (extraction of m-RNA, D
cleavage of NA with restriction enzymes, etc.) is performed using the method of Manianais et al. (Molecular Cloning, Col.
SprrngHarbor Laboratory
1982), the method of Berwin et al. (see DNA clo
ning, a Practical Approac.
h vol.
1、 p49. IRL、 0xford、 1935
年参照)を参照1:行った。1, p49. IRL, Oxford, 1935
Reference 1: I went.
マス、マウス・ミエローマ細胞株P3U1 (AT C
CCRL 1957)からmRNAを抽出し、ラムダフ
ァージgtlO(アマ−ジャム)を用いてcDNAライ
ブラリーを作製した。次にpBsF2R,236(K、
Yamasakiら、5cience、 241. p
825.1988年参照〉のインサートcDNAのFs
pl−Banl断片をプローブとして用い、7゜3X1
0’ クローンを以下の条件でスクリーニングした。プ
ラークをプロットするフィルターにはナイロンフィルタ
ーであるシーンスクリーンプラス(NEN社)を用いた
、ハイブリーダイゼーションは、1%SO3,LM N
aCj!、 0.05M Tris H1j! (p
H7,5)、5Xデンハルト溶液、20Off /−サ
ケ精子DNA存在下で、65℃で16時間行った。その
後、2 X5SC,1%SO3で、60℃にてフィルタ
ーを洗って非特異的にフィルターに結合したプローブを
分離させ、乾燥後、オートラジオグラフィーを行った。Trout, mouse myeloma cell line P3U1 (AT C
CCRL 1957) was extracted, and a cDNA library was created using lambda phage gtlO (Amar-Jam). Next, pBsF2R,236(K,
Yamasaki et al., 5science, 241. p
825.1988> insert cDNA Fs
Using the pl-Banl fragment as a probe, 7°3X1
0' clones were screened under the following conditions. A nylon filter, Scene Screen Plus (NEN), was used as a filter for plotting plaques. Hybridization was carried out using 1% SO3, LM N
aCj! , 0.05M Tris H1j! (p
H7,5), 5X Denhardt's solution, and 20Off/- salmon sperm DNA at 65°C for 16 hours. Thereafter, the filter was washed with 2×5SC, 1% SO3 at 60°C to separate the probe non-specifically bound to the filter, and after drying, autoradiography was performed.
なお、前記プラスミドpBsF2R,236を制限酵素
Xho Iで部分消火し、BSF 2レセプターをコー
ドする塩基配列をすべて含有するDNA断片を得、これ
を市販のベクターplBI76 (181社)のSal
1部位に挿入してプラスミドpIBIBsF2Rを作
製した。The plasmid pBsF2R, 236 was partially quenched with the restriction enzyme Xho I to obtain a DNA fragment containing the entire base sequence encoding the BSF 2 receptor, and this was transformed into a SalI fragment of the commercially available vector plBI76 (Company 181).
1 site to create plasmid pIBIBsF2R.
このプラスミドplBIBsF2Rを含有する大腸菌H
BIOI−plBIBsF2Rは、工業技術院微生物工
業技術研究所に微工研条寄第2232号(FBRM B
P−2232)として寄託されている。このプラスミド
から、常法に従って適当な制限酵素によりBSF 2レ
セプター蛋白質をコードするDNA断片を切り出し、本
発明で用いるプローブを作製することができる。E. coli H containing this plasmid plBIBsF2R
BIOI-plBIBsF2R was submitted to the Institute of Microbial Technology, Agency of Industrial Science and Technology, as part of the Microbiological Research Institute No. 2232 (FBRM B
P-2232). The probe used in the present invention can be prepared by cutting out a DNA fragment encoding the BSF 2 receptor protein from this plasmid using an appropriate restriction enzyme according to a conventional method.
このようにして単離された1つのクローンをラムダP
1 (lambda Pi) と名付けた、第1図にラ
ムダP1の制限酵母地図および対応するIL−6レセプ
ター蛋白質の模式図を示す。しかしながら、ラムダP1
めインサー) cDNAの塩基配列を決定した結果、ヒ
トーIL−6レセプターとの比較より、完全長のマウス
IL−6レセプターがコードされていないことが判明し
た。One clone thus isolated was used as lambda P
Figure 1 shows a schematic representation of the restriction yeast map of lambda P1 and the corresponding IL-6 receptor protein. However, lambda P1
As a result of determining the nucleotide sequence of the cDNA and comparing it with the human IL-6 receptor, it was found that the full-length mouse IL-6 receptor was not encoded.
そこで、完全長のマウスIL−6レセプターcDNAを
単離する目的で、上記方法でBALB/cの膵臓からm
RNAを抽出し、gtlOを用いてcDNAライブラリ
ーを作製した。プローブとして図1のプローブを用い、
通常の方法でスクリーニングした結果、1つのクローン
、ラムダ301 (Iambda301)を単離した、
ラムダ301のインサー) cDNAの塩基配列を決定
した結果、ヒトIL−6レセプターとの比較より、完全
長のマウスIL−6レセプターがコードされていること
が判明した。ヒ)IL−5レセプターとマウスIL−6
レセプターの相同性は、DNAレベルで69%、アミノ
酸レベルで54%であった。Therefore, in order to isolate full-length mouse IL-6 receptor cDNA, m
RNA was extracted and a cDNA library was created using gtlO. Using the probe in Figure 1 as a probe,
As a result of screening by conventional methods, one clone, Lambda301 (Iambda301), was isolated.
As a result of determining the base sequence of the cDNA (inser of lambda 301), it was found that the full-length mouse IL-6 receptor was encoded by comparison with the human IL-6 receptor. H) IL-5 receptor and mouse IL-6
The homology of the receptors was 69% at the DNA level and 54% at the amino acid level.
またラムダP1とラムダ301のそれぞれコードするマ
ウスIL−6レセプターcDNA塩基配列を比較したと
ころ、第1番目のメチオニンから第385番目のロイシ
ンまでをコードするDNA配列は一致したが、それ以降
のアミノ酸をコードするDNA配列は全く異なっていた
。さらに詳細な塩基配列の解析を行ったところ、ラムダ
P1の第386番目以降のアミノ酸をコードするDNA
配列は、1ntracysternal A−part
icle(IAP)遺伝子由来であることが判明した。Furthermore, when we compared the mouse IL-6 receptor cDNA base sequences encoded by lambda P1 and lambda 301, we found that the DNA sequences encoding from the 1st methionine to the 385th leucine matched, but the subsequent amino acids The encoding DNA sequences were completely different. Further detailed analysis of the base sequence revealed that the DNA encoding the amino acids starting from the 386th position of lambda P1
The sequence is 1ntracysternal A-part
icle (IAP) gene.
第1図に、ラムダ301の制限酵素地図および対応する
IL−6レセプター蛋白質の模式図を示す。FIG. 1 shows a restriction enzyme map of lambda 301 and a schematic diagram of the corresponding IL-6 receptor protein.
第2図に、ラムダ301のインサー)cDNAの塩基配
列および塩基配列から推定されるマウスIL−6レセプ
ターのアミノ酸配列を示す。FIG. 2 shows the nucleotide sequence of the inserter cDNA of lambda 301 and the amino acid sequence of the mouse IL-6 receptor deduced from the nucleotide sequence.
田賀らにより、ヒトIL−6レセプターの細胞外領域が
、IL−6との結合およびシグナルの伝達に必須で、細
胞内領域はこれらの目的には不要であることが示された
(Cell、 58. p573.1989年参照)。Taga et al. showed that the extracellular region of the human IL-6 receptor is essential for binding to IL-6 and transmitting signals, while the intracellular region is dispensable for these purposes (Cell, 58 (See p. 573, 1989).
単離したラムダ301のインサートcDNAが、マウス
tL−6レセプターのall)NAであること、および
、マウスIL−6レセプター蛋白質の細胞外領域のみが
IL−6との結合およびシグナルの伝達に必須であるこ
とを示す目的で、ラムダP1のインサ−)cDNA (
細胞外領域をコードするDNAはラムダ301と一致す
るが、膜貫通領域および細胞内領域はラムダ301 と
異なる。)、あるいはラムダ301のインサートC口N
^を細胞に導入し、マウスIL−6レセプターを細胞膜
上に発現させ、IL−6によるシグナル伝達を調べた。The isolated insert cDNA of lambda 301 is all) NA of the mouse tL-6 receptor, and only the extracellular region of the mouse IL-6 receptor protein is essential for binding to IL-6 and transmitting the signal. For the purpose of demonstrating that there is a lambda P1 inserter) cDNA (
The DNA encoding the extracellular region is identical to lambda301, but the transmembrane region and intracellular region are different from lambda301. ), or Lambda 301 insert C port N
^ was introduced into cells, mouse IL-6 receptor was expressed on the cell membrane, and signal transduction by IL-6 was investigated.
まず、アミノ酸コード領域を完全に含むラムダP1のE
coRI−EcoRI断片、およびラムダ301のBc
oRI−Kpnl断片をそれぞれ、ネオマインシ耐性遺
伝子を含む動物細胞発現ベクター、9M5G (C1L
akerら、Proc、natn、Acad、Sci、
U、S、A、、 84.p8459. 1987年参
照)に組み込んだ。次に、10%牛血清、50unit
s/mi7ペニシリン、50n/−ストレプトマイシン
、2mMグルタミン、50mM2−メルカプトエタノー
ル、10units/In!ヒトIL−6を含むRPM
11640培地で培養したヒ)IL−6依存性ヒトT細
胞株、KT −3(S、Shimizuら、Blood
72. p1826゜1988年参照)に、通常のエ
レクトロポーレーション法で、作製したベクターをそれ
ぞれ導入した。First, the E of lambda P1, which completely contains the amino acid coding region,
coRI-EcoRI fragment, and Bc of lambda 301
The oRI-Kpnl fragment was inserted into an animal cell expression vector, 9M5G (C1L
aker et al., Proc, natn, Acad, Sci,
U, S, A,, 84. p8459. (see 1987). Next, 10% bovine serum, 50 units
s/mi7 penicillin, 50n/-streptomycin, 2mM glutamine, 50mM 2-mercaptoethanol, 10 units/In! RPM containing human IL-6
IL-6-dependent human T cell line, KT-3 (S, Shimizu et al., Blood
72. p1826, 1988) by the usual electroporation method, the prepared vectors were introduced into each of the vectors.
750x/mj’の6418 (シグマ社)を含む上記
組成の培地でスクリーニングを行い、最終的に、2種類
の形質転換細胞を得た。Screening was performed using a medium with the above composition containing 6418 (Sigma) at 750x/mj', and finally two types of transformed cells were obtained.
この形質転換細胞2.5 XIO’を種々の濃度のマウ
ス・リコンビナントIL−6あるいは6 ng/rnl
のヒト・リコンビナントIL−6存在下で66時間培養
した。最後の6時間は、0.5μCiのトリチウム標識
チミジンでパルスし、細胞への取り込みをシンチレーシ
ョンカウンター(ベックマン社)で測定した。The transformed cells were treated with 2.5 XIO' at various concentrations of mouse recombinant IL-6 or 6 ng/rnl.
The cells were cultured for 66 hours in the presence of human recombinant IL-6. For the last 6 hours, the cells were pulsed with 0.5 μCi of tritium-labeled thymidine, and the uptake into the cells was measured using a scintillation counter (Beckman).
第3図から明らかなように、ラムダPl由来のマウスI
L−6レセプターが発現している細胞、および、ラムダ
301由来のマウスIL−6レセプターが発現している
細胞はマウス・リコンビナン)IL−6に濃度依存的に
増殖する。一方、形質転換されていないKT−3は、マ
ウス・リコンビナントIL−6に反応しない。また、3
種類の細胞ともヒト・リコンビナントIL−6には反応
する。これらの事実は、ラムダ301のインサートDN
AはマウスIL−6レセプターのcDNAであることを
証明するものであり、マウスIL−6レセプターはヒト
IL−6レセプターと同様に、細胞外領域がIL−6と
の結合およびシグナルの伝達に重要であり、細胞内領域
は他の配列に置き換えられることを示す。As is clear from Fig. 3, mouse I derived from lambda Pl
Cells expressing L-6 receptor and cells expressing mouse IL-6 receptor derived from lambda 301 proliferate in a concentration-dependent manner on mouse recombinant IL-6. On the other hand, untransformed KT-3 does not respond to mouse recombinant IL-6. Also, 3
All types of cells react with human recombinant IL-6. These facts indicate that the insert DN of Lambda 301
A proves that it is a cDNA of mouse IL-6 receptor, and like human IL-6 receptor, the extracellular region of mouse IL-6 receptor is important for binding with IL-6 and transmitting signals. , indicating that the intracellular region is replaced with another sequence.
本発明で提供されるマウスIL−6レセプターをコード
するDNA配列は、さらには該蛋白質を遺伝子工学的に
生産するための手段および方法により、自然状態では極
めて微量にしか生産されない該蛋白質を大量に生産する
ことが可能である。The DNA sequence encoding the mouse IL-6 receptor provided by the present invention can also be used to produce a large amount of the protein, which is produced in extremely small amounts in nature, by means and methods for producing the protein through genetic engineering. It is possible to produce.
また本発明により、マウスIL−6レセプターの生体内
での諸性質を解析および推定することが可能である。こ
のことはIL−6レセプターさらには生体の免疫機構等
の研究、またはそれらを用いた免疫疾患に対する治療薬
診断薬等の開発等に大きな意義をもつものである。Furthermore, according to the present invention, it is possible to analyze and estimate various properties of the mouse IL-6 receptor in vivo. This has great significance for research on IL-6 receptors and the immune system of living bodies, and for the development of therapeutic agents and diagnostic agents for immune diseases using them.
第1図は、ラムダP1およびラムダ301の制限酵素地
図および対応するIL−6レセプター蛋白質の模式図、
さらにはプローブ1の位置を示す。
Bは3am)If サイト、Eは!EC0RI サイト
、FはFOにIサイト、Hは旧ndIIIサイト、Kは
Kpnlサイト、PはPstlサイト、SはSmalサ
イトをそれぞれ示す。
また、SSはシグナル配列、ECは細胞外領域、TMは
膜貫通領域、Cは細胞内領域を示す。
第2図は、ラムダ301のインサートC口NA、すなわ
ちマウスIL−6レセプターをコードするDNA配列に
ついてその塩基配列を解析した結果、およびこのDNA
配列が発現された場合に生産される蛋白質すなわちマウ
スIL−6レセプターの推定されるアミノ酸配列を示す
。下線部分はN末端側の疎水性アミノ酸領域を示し、二
重下線部分はC末端側の疎水性アミノ酸領域を示す。
第3図は、ラムダP1由来のマウスIL−6レセプター
発現KT−3(○)、ラムダ301由来マウスIL−6
レセブタ一発現KT−3(・)、正常のKT−3(ロ)
の、種々の濃度のマウス・リコンビナントIL−6存在
下でのトリチウム標識チミジンの取り込みを示す。FIG. 1 shows restriction enzyme maps of lambda P1 and lambda 301 and a schematic diagram of the corresponding IL-6 receptor protein;
Furthermore, the position of probe 1 is shown. B is 3am) If site, E is! EC0RI site, F indicates FO I site, H indicates old ndIII site, K indicates Kpnl site, P indicates Pstl site, and S indicates Smal site. Further, SS represents a signal sequence, EC represents an extracellular region, TM represents a transmembrane region, and C represents an intracellular region. Figure 2 shows the results of nucleotide sequence analysis of the DNA sequence encoding the insert C-NA of lambda 301, that is, the mouse IL-6 receptor, and the results of this DNA sequence.
The deduced amino acid sequence of the protein produced when the sequence is expressed, namely the mouse IL-6 receptor, is shown. The underlined portion indicates the hydrophobic amino acid region on the N-terminal side, and the double underlined portion indicates the hydrophobic amino acid region on the C-terminal side. Figure 3 shows mouse IL-6 receptor expression KT-3 derived from lambda P1 (○), mouse IL-6 derived from lambda 301.
Receptor-expressing KT-3 (・), normal KT-3 (b)
Figure 3 shows the uptake of tritium-labeled thymidine in the presence of various concentrations of mouse recombinant IL-6.
Claims (1)
う。)と特異的に結合しうるマウス・インターロイキン
−6レセプター蛋白質。 2、下記の式( I ): (N末端) 【遺伝子配列があります】 (C末端) (ただし、式中Alaはアラニン、Argはアルギニン
、Asnはアスパラギン、Aspはアスパラギン酸、C
ysはシステイン、Glnはグルタミン、Gluはグル
タミン酸、Glyはグリシン、Hisはヒスチジン、I
leはイソロイシン、Leuはロイシン、Lysはリジ
ン、Metはメチオニン、Pheはフェニルアラニン、
Proはプロリン、Serはセリン、Thrはトレオニ
ン、Trpはトリプトファン、Tyrはチロシン、そし
てValはバリン残基を示す。)で表されるアミノ酸配
列を有するか、あるいはこのアミノ酸配列中の1個また
は複数個のアミノ酸残基が他のアミノ酸残基により置換
されているかまたは欠失もしくは付加されているアミノ
酸配列を有し、かつマウスIL−6と特異的に結合する
能力を保持していることを特徴とする請求項1に記載の
マウスIL−6レセプター蛋白質。 3、マウスIL−6レセプター蛋白質をコードするDN
A。 4、請求項2に記載のいずれかのアミノ酸配列を有する
マウスIL−6レセプター蛋白質をコードするDNA。 5、下記の式(II): (5^−) 【遺伝子配列があります】 (3^−) (ただしAはアデニン、Gはグアニン、Cはシトシン、
Tはチミンを有するヌクレオシドを示す)で表される請
求項3に記載のDNA。 6、前記DNAを発現させうるDNA配列と読取り可能
に結合していることを特徴とする請求項3に記載のDN
A。 7、組換微生物または培養細胞中で請求項3に記載のD
NA配列を発現しうる複製可能な発現ベクター。 8、請求項7に記載の発現ベクターにより形質転換され
た微生物または培養細胞においてマウスIL−6レセプ
ター蛋白質をコードするDNA配列を発現させ、該蛋白
質を生産させることを特徴とするマウスIL−6レセプ
ター蛋白質の生産方法。[Scope of Claims] 1. A mouse interleukin-6 receptor protein that can specifically bind to mouse interleukin-6 (hereinafter referred to as IL-6). 2. The following formula (I): (N-terminus) [There is a gene sequence] (C-terminus) (However, in the formula, Ala is alanine, Arg is arginine, Asn is asparagine, Asp is aspartic acid, C
ys is cysteine, Gln is glutamine, Glu is glutamic acid, Gly is glycine, His is histidine, I
le is isoleucine, Leu is leucine, Lys is lysine, Met is methionine, Phe is phenylalanine,
Pro represents proline, Ser represents serine, Thr represents threonine, Trp represents tryptophan, Tyr represents tyrosine, and Val represents valine. ), or has an amino acid sequence in which one or more amino acid residues in this amino acid sequence are substituted with other amino acid residues, or are deleted or added. 2. The mouse IL-6 receptor protein according to claim 1, which retains the ability to specifically bind to mouse IL-6. 3. DN encoding mouse IL-6 receptor protein
A. 4. DNA encoding a mouse IL-6 receptor protein having the amino acid sequence according to claim 2. 5. Formula (II) below: (5^-) [There is a gene sequence] (3^-) (A is adenine, G is guanine, C is cytosine,
4. The DNA according to claim 3, wherein T represents a nucleoside having thymine. 6. The DNA according to claim 3, wherein the DNA is readably linked to a DNA sequence capable of expressing the DNA.
A. 7. D according to claim 3 in a recombinant microorganism or cultured cells.
A replicable expression vector capable of expressing an NA sequence. 8. A mouse IL-6 receptor, characterized in that the protein is produced by expressing a DNA sequence encoding the mouse IL-6 receptor protein in a microorganism or cultured cell transformed with the expression vector according to claim 7. Protein production methods.
Priority Applications (1)
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JP1292230A JPH03155795A (en) | 1989-11-13 | 1989-11-13 | Mouse-interleukin-6 receptor protein |
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Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP1292230A JPH03155795A (en) | 1989-11-13 | 1989-11-13 | Mouse-interleukin-6 receptor protein |
Publications (1)
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JPH03155795A true JPH03155795A (en) | 1991-07-03 |
Family
ID=17779174
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