MAP dendrimer elicits antibodies for detecting rat and mouse GH-binding proteins

被引:5
作者
Aguilar, Roberto M. [2 ]
Talamantes, Frank J. [3 ]
Bustamante, Juan J. [4 ]
Munoz, Jesus [6 ]
Trevino, Lisa R. [5 ]
Martinez, Andrew O. [1 ]
Haro, Luis S. [1 ]
机构
[1] Univ Texas San Antonio, Dept Biol, San Antonio, TX 78249 USA
[2] Univ Calif Irvine, Reeve Irvine Res Ctr, Irvine, CA 92697 USA
[3] Univ Calif Santa Cruz, Dept Biol, Santa Cruz, CA 95060 USA
[4] Texas A&M Hlth Sci Ctr, Irma Lerma Rangel Coll Pharm, Kingsville, TX 78363 USA
[5] St Jude Childrens Res Hosp, Dept Pharmaceut Sci, Memphis, TN 38105 USA
[6] Univ Texas San Antonio, Dept Chem, San Antonio, TX 78249 USA
基金
美国国家卫生研究院;
关键词
MAP dendrimer; antipeptide polyclonal antisera; GH-binding protein; GH receptor; GH; GROWTH-HORMONE RECEPTOR; SYNTHETIC PEPTIDE VACCINE; GLYCOPEPTIDE DENDRIMERS; TRANSGENIC MICE; KNOCKOUT MICE; SERUM; EXPRESSION; SYSTEM; DESIGN; LIVER;
D O I
10.1002/psc.1096
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The membrane-bound rat GH-R and an alternatively spliced isoform, the soluble rat GH-BP, are comprised of identical N-terminal GH-binding domains; however, their C-terminal sequences differ. Immunological reagents are needed to distinguish between the two isoforms in order to understand their respective roles in mediating the actions of GH. Accordingly, a tetravalent MAP dendrimer with four identical branches of a C-terminal peptide sequence of the rat GH-BP (GH-BP263-279) was synthesized and used as an immunogen in rabbits. Solid-phase peptide synthesis of four GH-BP263-279 segments onto a tetravalent Lys(2)-Lys beta-Ala-OH core peptide was carried out using Fmoc chemistry. The mass of the RP-HPLC-purified synthetic product, 8398 Da, determined by ESI-MS, was identical to expected mass. Three anti-rat GH-BP263-279 MAP antisera, BETO-8039, BETO-8040, and BETO-8041, at dilutions of 10(-3), recognized both the rat GH-BP263-279 MAP and recombinant mouse GH-BP with ED(50)s within a range of 5-10 fmol, but did not cross-react with BSA in dot blot analyses. BETO-8041 antisera (10-3 dilution) recognized GH-BPs of rat serum and liver having M(r)s ranging from 35 to 130 kDa, but did not recognize full-length rat GH-Rs. The antisera also detected recombinant mouse GH-BPs. In summary, the tetravalent rat GH-BP263-279 MAP dendrimer served as an effective immunogenic antigen in eliciting high titer antisera specific for the C-termini of both rat and mouse GH-BPs. The antisera will facilitate studies aimed at improving our understanding of the biology of GH-BPs. Copyright (c) 2008 European Peptide Society and John Wiley & Sons, Ltd.
引用
收藏
页码:78 / 88
页数:11
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