In order to help uncover the basis of the strong biotin-avidin binding and to identify any contributory biotin-tryptophan interaction, we have synthesized model compounds, Biot-C3-Ind (4a) and Biot-C4-Ind (4b), having trimethylene and tetramethylene chains between the biotin ring system and indole, by starting with (+)-biotin and (+)-homobiotin. The UV spectroscopic effect of the proximate biotin moiety in Biot-C3-Ind compared with 3-propylindole, both in cyclohexane (with 1% ethanol), is similar to the effect of biotin bound to avidin. Interaction of the biotin ring system and indole was also evidenced by a red shift in λem in comparing the fluorescence emission of 3-propylindole with that of Biot-C3-Ind or Biot-C4-Ind in cyclohexane (1 or 0.1% ethanol), but the proximate biotin ring system caused no quenching of fluorescence. We have found no evidence of strong interaction between the biotin ring system and the indole of tryptophan such as would help account for the magnitude of the avidin-biotin association constant. The diastereomeric sulfoxides of Biot-C3-Ind were prepared, and the stereochemistry of each was established. 1H NMR spectral comparisons with the methyl esters of (+)-biotin and its two sulfoxides confirmed the stereochemical assignments, as did X-ray single-crystal analysis (the sequel). With the 1H NMR chemical shift assignments and the X-ray-confirmed stereochemistry, we have developed correlations with 13C NMR chemical shifts that should prove useful in assigning stereochemistry to other asymmetric sulfoxides. A quantitative study of the N-bromosuccinimide oxidation of model compounds and of avidin, biotin, and the avidin-biotin complex established the relative rates of oxidation and the stereochemistry of oxidation of the biotin moiety in various environments. The protection of the tryptophans in the avidin-biotin complex against NBS oxidation is due only partially to consumption of the oxidizing agent by the bound biotin. Biotin in the avidin-biotin complex is oxidized by aqueous NBS, yet it is more protected than free biotin in aqueous solution. Oxidation of biotin in the avidin-biotin complex yields predominantly the α-sulfoxide, indicating steric limitation of the approach of oxidant to bound biotin.
|Original language||English (US)|
|Number of pages||10|
|Journal||Journal of the American Chemical Society|
|State||Published - 1979|
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