High-Level Expression of TGF-beta 2 and the TGF-beta 2(414) Precursor in Chinese Hamster Ovary Cells

被引：22

作者：

Madisen, Linda ^{[1
]}

Lioubin, Mario N. ^{[1
]}

Marquardt, Hans ^{[1
]}

Purchio, A. F. ^{[1
]}

机构：

[1] ONCOGEN, 3005 1st Ave, Seattle, WA 98121 USA

来源：

GROWTH FACTORS | 1990年 / 3卷 / 02期

关键词：

TGF-beta; 2; in vitro expression; immunoblotting; protein sequencing;

D O I：

10.3109/08977199009108275

中图分类号：

Q2 [细胞生物学];

学科分类号：

071009 ; 090102 ;

摘要：

Chinese hamster ovary (CHO) clones secreting high levels of transforming growth factor-beta 2 (TGF-beta 2) were obtained after transfection with a cDNA clone coding for the 414-amino acid TGF-beta 2 precursor and subsequent amplification with methotrexate. The TGF-beta 2 was secreted in a latent form since acidification was necessary for detection of maximal levels of bioactivity. Amino- and carboxy-terminal sequencing of purified recombinant TGF-beta 2 indicated that correct processing of mature TGF-beta 2 had occurred. In addition to mature TGF-beta 2, the recombinant CHO clones secreted larger proteins having molecular weights of 85, 105, and 130 kD, which consisted of both mature and pro-region sequences when analyzed by immunoblotting using site-specific anti-peptide antibodies. Analysis of serumand cell-free media from recombinant CHO cells metabolically labeled with [H-3] glucosamine and [P-32] orthophosphateindicated that pro-TGF-beta 2 was glycosylated and phosphorylated. Two-dimensional electrophoretic analysis of acid hydrolysates showed that the P-32 was incorporated into mannose-6-phosphate.

引用

页码：129 / 138

页数：10

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