کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
10769102 | 1050819 | 2005 | 5 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Engineering of Kex2 variants exhibiting altered substrate specificity
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کلمات کلیدی
موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
زیست شیمی
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
Engineering of secreted protease variants exhibiting altered substrate specificity is a challenging task because effective screening methods for the desired property are not available yet. In this study, we sought to obtain variants of Kex2, a yeast Golgi protease, which exhibit altered P2 specificity. We first randomly mutated three Asp residues (D176, D210, and D211) that constitute the S2 pocket of Kex2 and then isolated from the resulting library Kex2 variants that preferred substrates with Met (poorly preferred by wild type Kex2) at the P2 position using a yeast-based screening method. The Kex2 variants isolated from this initial screening were further tested against various substrate sequences. Four out of the 16 isolated Kex2 variants showed greater preference for Met than for Lys (preferred by the wild-type Kex2) at the P2 position. We therefore suggest that our method might serve as an efficient tool for engineering and directing the evolution of secreted proteases.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Biochemical and Biophysical Research Communications - Volume 337, Issue 4, 2 December 2005, Pages 1102-1106
Journal: Biochemical and Biophysical Research Communications - Volume 337, Issue 4, 2 December 2005, Pages 1102-1106
نویسندگان
Hye-Eun Han, Seong-Hwan Rho, Yong Jae Lee, Woo Jin Park,