Design, production, and characterization of a monomeric streptavidin and its application for affinity purification of biotinylated proteins

Qureshi, Mohammad Hassan; Wong, Sui‐Lam

doi:10.1016/s1046-5928(02)00021-9

Cited by 48 publications

(36 citation statements)

References 21 publications

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“…** Disruption of protein quaternary structure, however, often results in lower catalytic activity, reduced binding affinity, and decreased solubility and stability. However, this paper and other work (6)(7)(8)(9)(10) clearly demonstrate that soluble functional dimeric and monomeric streptavidins and avidins can be engineered.…”

mentioning

confidence: 66%

Engineered single-chain dimeric streptavidins with an unexpected strong preference for biotin-4-fluorescein

Aslan

Yu²,

Mohr³

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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mentioning

confidence: 66%

Engineered single-chain dimeric streptavidins with an unexpected strong preference for biotin-4-fluorescein

Aslan

Yu²,

Mohr³

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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“…It was also stated in a subsequent study by Stayton's group that the structure closely parallels a key intermediate observed in a simulated dissociation pathway of biotin from streptavidin [53]. The biotin-binding residues Ser45, Thr90 and Asp128 of streptavidin have been converted separately, in combinations of two, or of all three to alanines [54,55]. These mutants showed a wide spectrum of affinities toward biotin (K d = 10 -6 -10 -11 M).…”

Section: Binding-site Mutantsmentioning

confidence: 68%

“…Qureshi and Wong were able to turn streptavidin into a soluble monomer by converting two biotin-binding residues, Thr90 and Asp128, simultaneously into alanines [55]. They reported the K d of the resultant mutant to be 1.3 × 10 -8 M toward biotin and showed successful utilization of the immobilized mutant in the purification of biotinylated cytochrome C from a bacterial extract, and then its mild elution by desthiobiotin.…”

Section: Interface Mutantsmentioning

confidence: 99%

“…These monomeric streptavidins, based on interface residue modifications, showed approximately four times weaker affinity toward biotin when compared with monomeric streptavidin, where monomerization was achieved by mutating biotin-binding residues T90A and D128A [54,55,67]. In the same study Wu and Wong [67] also constructed a streptavidin mutant where they mimicked our strategy to monomerize avidin.…”

Section: Interface Mutantsmentioning

confidence: 99%

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Genetically engineered avidins and streptavidins

Laitinen

Hytönen

Nordlund

et al. 2006

Cell. Mol. Life Sci.

295

197

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Chicken avidin and bacterial streptavidin, (strept)avidin, are proteins widely utilized in a number of applications in life science, ranging from purification and labeling techniques to diagnostics, and from targeted drug delivery to nanotechnology. (Strept)avidin-biotin technology relies on the extremely tight and specific affinity between (strept)avidin and biotin (dissociation constant, K(d) approximately 10(-14)-10(-16) M). (Strept)avidins are also exceptionally stable proteins. To study their ligand binding and stability characteristics, the two proteins have been extensively modified both chemically and genetically. There are excellent accounts of this technology and chemically modified (strept)avidins, but no comprehensive reviews exist concerning genetically engineered (strept)avidins. To fill this gap, we here go through the genetically engineered (strept)avidins, summarizing how these constructs were designed and how they have improved our understanding of the structural and functional characteristics of these proteins, and the benefits they have provided for (strept)avidin-biotin technology.

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“…It should be possible to maintain the high affinity toward biotin in two of the binding sites while modifying the affinity in the other two sites as desired. One option could be to make two of the binding sites reversible, which would enable bound materials to be mildly detached by free biotin (11,14,32,33). Another procedure might be to modify half of the binding sites with smart polymer conjugates able to respond to changes in pH, light intensity, and temperature (34,35).…”

Section: Fig 1 Description Of the Avidin Mutantsmentioning

confidence: 99%

Construction of a Dual Chain Pseudotetrameric Chicken Avidin by Combining Two Circularly Permuted Avidins

Nordlund

Laitinen

Hytönen

et al. 2004

Journal of Biological Chemistry

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Two distinct circularly permuted forms of chicken avidin were designed with the aim of constructing a fusion avidin containing two biotin-binding sites in one polypeptide. The old N and C termini of wild-type avidin were connected to each other via a glycine/serine-rich linker, and the new termini were introduced into two different loops. This enabled the creation of the desired fusion construct using a short linker peptide between the two different circularly permuted subunits. The circularly permuted avidins (circularly permuted avidin 5 3 4 and circularly permuted avidin 6 3 5) and their fusion, pseudotetrameric dual chain avidin, were biologically active, i.e. showed biotin binding, and also displayed structural characteristics similar to those of wild-type avidin. Dual chain avidin facilitates the development of dual affinity avidins by allowing adjustment of the ligand-binding properties in half of the binding sites independent of the other half. In addition, the subunit fusion strategy described in this study can be used, where applicable, to modify oligomeric proteins in general.

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Design, production, and characterization of a monomeric streptavidin and its application for affinity purification of biotinylated proteins

Cited by 48 publications

References 21 publications

Engineered single-chain dimeric streptavidins with an unexpected strong preference for biotin-4-fluorescein

Engineered single-chain dimeric streptavidins with an unexpected strong preference for biotin-4-fluorescein

Genetically engineered avidins and streptavidins

Construction of a Dual Chain Pseudotetrameric Chicken Avidin by Combining Two Circularly Permuted Avidins

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