2CAM

AVIDIN MUTANT (K3E,K9E,R26D,R124L)

2CAM の概要

• エントリーDOI: 10.2210/pdb2cam/pdb
• 分子名称: AVIDIN (2 entities in total)
• 機能のキーワード: avidin, biotin binding protein, calycins, up-and-down beta barrel, glycoprotein
• 由来する生物種: Gallus gallus (chicken)
• 細胞内の位置: Secreted: P02701
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 28409.57

構造登録者

Rosano, C.,Arosio, P.,Bolognesi, M. (登録日: 1998-03-27, 公開日: 1998-07-15, 最終更新日: 2024-10-09)

主引用文献

Nardone, E.,Rosano, C.,Santambrogio, P.,Curnis, F.,Corti, A.,Magni, F.,Siccardi, A.G.,Paganelli, G.,Losso, R.,Apreda, B.,Bolognesi, M.,Sidoli, A.,Arosio, P.
Biochemical characterization and crystal structure of a recombinant hen avidin and its acidic mutant expressed in Escherichia coli.
Eur.J.Biochem., 256:453-460, 1998

PubMed Abstract: The mature hen avidin encoded by a synthetic cDNA was expressed in Escherichia coli in an insoluble form. After resolubilization, renaturation and purification, a recovery of about 20 mg/l cell culture was obtained. ELISA assays indicated no apparent differences in biotin binding between the natural and recombinant avidins. In addition, an acidic avidin mutant, bearing the substitutions Lys3-->Glu, Lys9--> Glu, Arg26-->Asp and Arg124-->Leu of four exposed basic residues, was produced. The protein, expressed and renatured as wild-type avidin, showed unaltered biotin-binding activity. The acidic pI (approximately 5.5) and lack of aggregation of the mutant allowed easy electrophoretic analysis under non-denaturing conditions of the protein alone and of its complexes with biotin, biotinylated transferrin or peroxidase. Analysis of the sera from sensitized subjects revealed that the avidin mutant has altered antigenicity. Both recombinant avidins were crystallized and the three-dimensional structures solved by molecular replacement and refined to 0.22 nm resolution. The three-dimensional structures of the two recombinant molecules, in the absence of biotin and of glycosylation, are fully comparable with those of the natural hen avidin previously reported.

PubMed: 9760187
DOI: 10.1046/j.1432-1327.1998.2560453.x

実験手法

X-RAY DIFFRACTION (2.2 Å)