Chicken avidin-related protein 4/5 shows superior thermal stability when compared with avidin while retaining high affinity to biotin

Hytönen, Thomas Nyholm, Pentikäinen, Vaarno, Porkka, Nordlund, Mark S Johnson, J Peter Slotte, Laitinen, Kulomaa

Research output: Contribution to journalArticleScientificpeer-review

44 Citations (Scopus)

Abstract

The protein chicken avidin is a commonly used tool in various applications. The avidin gene belongs to a gene family that also includes seven other members known as the avidin-related genes (AVR). We report here on the extremely high thermal stability and functional characteristics of avidin-related protein AVR4/5, a member of the avidin protein family. The thermal stability characteristics of AVR4/5 were examined using a differential scanning calorimeter, microparticle analysis, and a microplate assay. Its biotin-binding properties were studied using an isothermal calorimeter and IAsys optical biosensor. According to these analyses, in the absence of biotin AVR4/5 is clearly more stable (T(m) = 107.4 +/- 0.3 degrees C) than avidin (T(m) = 83.5 +/- 0.1 degrees C) or bacterial streptavidin (T(m) = 75.5 degrees C). AVR4/5 also exhibits a high affinity for biotin (K(d) approximately 3.6 x 10(-14) m) comparable to that of avidin and streptavidin (K(d) approximately 10(-15) m). Molecular modeling and site-directed mutagenesis were used to study the molecular details behind the observed high thermostability. The results indicate that AVR4/5 and its mutants have high potential as new improved tools for applications where exceptionally high stability and tight biotin binding are needed.
Original languageUndefined/Unknown
Pages (from-to)9337–9343
JournalJournal of Biological Chemistry
Volume279
Issue number10
DOIs
Publication statusPublished - 2004
MoE publication typeA1 Journal article-refereed

Cite this