Selection of a biotin protein ligase by phage display using a combination of in vitro selection and in vivo enzymatic activity

被引：2

作者：

Iwamoto, Masahiro ^{[2
]}

Taki, Takashi ^{[2
]}

Fujita, Satoshi ^{[1
,2
]}

机构：

[1] Natl Inst Adv Ind Sci & Technol, Res Inst Cell Engn, Koto Ku, Tokyo 1350064, Japan

[2] Univ Tokyo, Grad Sch Engn, Dept Chem & Biotechnol, Tokyo 1138656, Japan

来源：

JOURNAL OF BIOSCIENCE AND BIOENGINEERING | 2009年 / 107卷 / 03期

基金：

日本学术振兴会;

关键词：

Phage display; Biotin protein ligase; Screening; Neurotensin; Random Library; BREAST-CANCER; NEUROTENSIN; ACTIVATION;

D O I：

10.1016/j.jbiosc.2008.11.009

中图分类号：

Q81 [生物工程学（生物技术）]; Q93 [微生物学];

学科分类号：

071005 ; 0836 ; 090102 ; 100705 ;

摘要：

We report the development of a phage display-based method for the direct selection of enzymatic activity by a combination of selection in vitro and an assay of enzymatic activity in vivo. We describe the selection of a biotin protein ligase (BPL) that specifically recognizes neurotensin as a substrate, as an example of the utility of our method. We constructed an enzyme library with a diversity of 8.8x10(6) by insertion of four randomized regions into a BPL backbone and used it for the selection of variant BPLs. We obtained an active BPL, which was identical to wild-type BPL, using the combination of selection in vitro and an assay of enzymatic activity in vivo. Our results indicate that our method is suitable for the selection of enzymes with specific functions of interest. (C) 2008. The Society for Biotechnology, Japan. All rights reserved.

引用

页码：230 / 234

页数：5

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