BETADOUBLET - DE-NOVO DESIGN, SYNTHESIS, AND CHARACTERIZATION OF A BETA-SANDWICH PROTEIN

被引:150
作者
QUINN, TP
TWEEDY, NB
WILLIAMS, RW
RICHARDSON, JS
RICHARDSON, DC
机构
[1] DUKE UNIV, DEPT BIOCHEM, DURHAM, NC 27710 USA
[2] UNIFORMED SERV UNIV HLTH SCI, DEPT BIOCHEM, BETHESDA, MD 20814 USA
关键词
PROTEIN DESIGN; BETA-STRUCTURE; MOLECULAR MODELING; CIRCULAR DICHROISM; THERMOSTABILITY;
D O I
10.1073/pnas.91.19.8747
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
How an amino acid sequence encodes the information necessary for a protein to adopt a unique tertiary structure remains unresolved. We are addressing this problem by designing ''from scratch'' protein molecules that will adopt predetermined three-dimensional structures. Based on this strategy, two identical four-stranded beta-sheets were designed to dimerize and form a beta-sandwich protein, called betadoublet. A synthetic gene encoding half the beta-sandwich protein was expressed in Escherichia coli, and the protein was purified to homogeneity. Biophysical characterization of betadoublet in aqueous solution demonstrated that the disulfide formed between the two sheets and that the dimer was a compact unaggregated globular protein, consisting predominantly of beta-sheet and stable to thermal denaturation. It has some backbone amide protons whose exchange is slow enough to be measured by NMR but binds more of the dye 1-anilinonaphthalene-8-sulfonate than a well-folded protein.
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页码:8747 / 8751
页数:5
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