Construction of chimeric dual-chain avidin by tandem fusion of the related avidins

Tiina A. Riihimäki, Sampo Kukkurainen, Suvi Varjonen, Jarno Hörhä, Thomas K.M. Nyholm, Markku S. Kulomaa, Vesa P. Hytönen (Corresponding Author)

Research output: Contribution to journalArticleScientificpeer-review

4 Citations (Scopus)

Abstract

Background

Avidin is a chicken egg-white protein with high affinity to vitamin H, also known as D-biotin. Many applications in life science research are based on this strong interaction. Avidin is a homotetrameric protein, which promotes its modification to symmetrical entities. Dual-chain avidin, a genetically engineered avidin form, has two circularly permuted chicken avidin monomers that are tandem-fused into one polypeptide chain. This form of avidin enables independent modification of the two domains, including the two biotin-binding pockets; however, decreased yields in protein production, compared to wt avidin, and complicated genetic manipulation of two highly similar DNA sequences in the tandem gene have limited the use of dual-chain avidin in biotechnological applications.

Principal Findings

To overcome challenges associated with the original dual-chain avidin, we developed chimeric dual-chain avidin, which is a tandem fusion of avidin and avidin-related protein 4 (AVR4), another member of the chicken avidin gene family. We observed an increase in protein production and better thermal stability, compared with the original dual-chain avidin. Additionally, PCR amplification of the hybrid gene was more efficient, thus enabling more convenient and straightforward modification of the dual-chain avidin. When studied closer, the generated chimeric dual-chain avidin showed biphasic biotin dissociation.

Significance

The improved dual-chain avidin introduced here increases its potential for future applications. This molecule offers a valuable base for developing bi-functional avidin tools for bioseparation, carrier proteins, and nanoscale adapters. Additionally, this strategy could be helpful when generating hetero-oligomers from other oligomeric proteins with high structural similarity.

Original languageEnglish
Article numbere20535
Number of pages10
JournalPLoS ONE
Volume6
Issue number5
DOIs
Publication statusPublished - 2011
MoE publication typeA1 Journal article-refereed

Fingerprint

avidin
Avidin
Fusion reactions
biotin
Biotin
Chickens
proteins
Genes
Proteins
chickens
Egg Proteins
genes
Biological Science Disciplines
Gene Amplification
chicken eggs
DNA sequences
transport proteins
egg albumen

Cite this

Riihimäki, T. A., Kukkurainen, S., Varjonen, S., Hörhä, J., Nyholm, T. K. M., Kulomaa, M. S., & Hytönen, V. P. (2011). Construction of chimeric dual-chain avidin by tandem fusion of the related avidins. PLoS ONE, 6(5), [e20535]. https://doi.org/10.1371/journal.pone.0020535
Riihimäki, Tiina A. ; Kukkurainen, Sampo ; Varjonen, Suvi ; Hörhä, Jarno ; Nyholm, Thomas K.M. ; Kulomaa, Markku S. ; Hytönen, Vesa P. / Construction of chimeric dual-chain avidin by tandem fusion of the related avidins. In: PLoS ONE. 2011 ; Vol. 6, No. 5.
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abstract = "BackgroundAvidin is a chicken egg-white protein with high affinity to vitamin H, also known as D-biotin. Many applications in life science research are based on this strong interaction. Avidin is a homotetrameric protein, which promotes its modification to symmetrical entities. Dual-chain avidin, a genetically engineered avidin form, has two circularly permuted chicken avidin monomers that are tandem-fused into one polypeptide chain. This form of avidin enables independent modification of the two domains, including the two biotin-binding pockets; however, decreased yields in protein production, compared to wt avidin, and complicated genetic manipulation of two highly similar DNA sequences in the tandem gene have limited the use of dual-chain avidin in biotechnological applications.Principal FindingsTo overcome challenges associated with the original dual-chain avidin, we developed chimeric dual-chain avidin, which is a tandem fusion of avidin and avidin-related protein 4 (AVR4), another member of the chicken avidin gene family. We observed an increase in protein production and better thermal stability, compared with the original dual-chain avidin. Additionally, PCR amplification of the hybrid gene was more efficient, thus enabling more convenient and straightforward modification of the dual-chain avidin. When studied closer, the generated chimeric dual-chain avidin showed biphasic biotin dissociation.SignificanceThe improved dual-chain avidin introduced here increases its potential for future applications. This molecule offers a valuable base for developing bi-functional avidin tools for bioseparation, carrier proteins, and nanoscale adapters. Additionally, this strategy could be helpful when generating hetero-oligomers from other oligomeric proteins with high structural similarity.",
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Riihimäki, TA, Kukkurainen, S, Varjonen, S, Hörhä, J, Nyholm, TKM, Kulomaa, MS & Hytönen, VP 2011, 'Construction of chimeric dual-chain avidin by tandem fusion of the related avidins', PLoS ONE, vol. 6, no. 5, e20535. https://doi.org/10.1371/journal.pone.0020535

Construction of chimeric dual-chain avidin by tandem fusion of the related avidins. / Riihimäki, Tiina A.; Kukkurainen, Sampo; Varjonen, Suvi; Hörhä, Jarno; Nyholm, Thomas K.M.; Kulomaa, Markku S.; Hytönen, Vesa P. (Corresponding Author).

In: PLoS ONE, Vol. 6, No. 5, e20535, 2011.

Research output: Contribution to journalArticleScientificpeer-review

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AU - Riihimäki, Tiina A.

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AU - Nyholm, Thomas K.M.

AU - Kulomaa, Markku S.

AU - Hytönen, Vesa P.

PY - 2011

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N2 - BackgroundAvidin is a chicken egg-white protein with high affinity to vitamin H, also known as D-biotin. Many applications in life science research are based on this strong interaction. Avidin is a homotetrameric protein, which promotes its modification to symmetrical entities. Dual-chain avidin, a genetically engineered avidin form, has two circularly permuted chicken avidin monomers that are tandem-fused into one polypeptide chain. This form of avidin enables independent modification of the two domains, including the two biotin-binding pockets; however, decreased yields in protein production, compared to wt avidin, and complicated genetic manipulation of two highly similar DNA sequences in the tandem gene have limited the use of dual-chain avidin in biotechnological applications.Principal FindingsTo overcome challenges associated with the original dual-chain avidin, we developed chimeric dual-chain avidin, which is a tandem fusion of avidin and avidin-related protein 4 (AVR4), another member of the chicken avidin gene family. We observed an increase in protein production and better thermal stability, compared with the original dual-chain avidin. Additionally, PCR amplification of the hybrid gene was more efficient, thus enabling more convenient and straightforward modification of the dual-chain avidin. When studied closer, the generated chimeric dual-chain avidin showed biphasic biotin dissociation.SignificanceThe improved dual-chain avidin introduced here increases its potential for future applications. This molecule offers a valuable base for developing bi-functional avidin tools for bioseparation, carrier proteins, and nanoscale adapters. Additionally, this strategy could be helpful when generating hetero-oligomers from other oligomeric proteins with high structural similarity.

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Riihimäki TA, Kukkurainen S, Varjonen S, Hörhä J, Nyholm TKM, Kulomaa MS et al. Construction of chimeric dual-chain avidin by tandem fusion of the related avidins. PLoS ONE. 2011;6(5). e20535. https://doi.org/10.1371/journal.pone.0020535