کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
2020408 | 1542341 | 2014 | 7 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Purification of recombinant nacre-associated mineralization protein AP7 fused with maltose-binding protein
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کلمات کلیدی
موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
زیست شیمی
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چکیده انگلیسی
Formation of biominerals often involves specific proteins that modulate the process of matrix assembly, nucleation, and crystal growth. AP7 is an aragonite-associated protein of 7 kDa and is intrinsically disordered. The structural disorder of AP7 makes it very difficult to express in Escherchia coli. In this work, we report the first successful expression and purification of recombinant AP7 using the maltose-binding protein (MBP) fusion approach. We obtain a high-yield production of recombinant MBP-AP7 protein in E. coli (â¼60 mg/L). We also establish an efficient protocol to remove the MBP fusion protein by Factor Xa, followed by purification using size-exclusion chromatography. Characterization of the recombinant AP7 protein has been carried out using MALDI-TOF, peptide mass fingerprinting, and circular dichroism (CD). The mass data confirm that the purified recombinant protein is AP7. The CD data suggest that the recombinant AP7 protein exists as partially disordered structure at neutral pH. The calcium carbonate precipitation assay shows that both MBP-AP7 and AP7 exhibit morphological modification on calcite crystallites. The co-precipitation of MBP-tagged AP7 derivatives and calcium carbonate generate different types of AP7 composite calcite and vaterite crystals. This system should be helpful to establish a model for understanding the structure/function relationship between the protein and inorganic mineral interaction.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Protein Expression and Purification - Volume 100, August 2014, Pages 26-32
Journal: Protein Expression and Purification - Volume 100, August 2014, Pages 26-32
نویسندگان
Yu-Chieh Huang, Hsun-Hui Chang, Yun Mou, Peter Chi, Jerry Chun Chung Chan, Shih-Chi Luo,