@article{Massey19866Thio-8234,
  year={1986},
  doi={10.1021/bi00372a046},
  title={6-Thiocyanatoflavins and 6-mercaptoflavins as active-site probes of flavoproteins},
  number={24},
  volume={25},
  issn={0006-2960},
  journal={Biochemistry},
  pages={8103--8112},
  author={Massey, Vincent and Ghisla, Sandro and Yagi, Kunio}
}

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    <dc:contributor>Ghisla, Sandro</dc:contributor>
    <dc:creator>Massey, Vincent</dc:creator>
    <dc:creator>Ghisla, Sandro</dc:creator>
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    <dcterms:abstract xml:lang="eng">6-Thiocyanatoflavins have been found to be susceptible to nucleophilic displacement reactions with sulfite and thiols, yielding respectively the 6-S-S03--flavin and 6-mercaptoflavin, with rate constants at pH 7.0, 20°C, of 55 M-1 min-1 for sulfite and 1000 M-1 min-1 for dithiothreitol. The 6-SCN-flavin binds tightly to riboflavin-binding protein as the riboflavin derivative, to apoflavodoxin, apo-lactate oxidase, and apo-Old Yellow Enzyme as the FMN derivative, and to apo-D-amino acid oxidase as the FAD derivative. The riboflavin-binding protein derivative is inaccessible to dithiothreitol attack, and the lactate oxidase and D-amino acid oxidase derivatives show only limited accessibility. However, the flavodoxin and Old Yellow Enzyme derivatives react readily with dithiothreitol, indicating that the flavin 6-position is exposed to solvent in these proteins. The lactate oxidase and D-amino acid oxidase derivatives convert slowly but spontaneously to the 6-mercaptoflavin enzyme forms in the absence of any added thiol, indicating the presence of a thiol residue in the flavin binding site of these proteins. The reaction rates have been investigated of 6-mercaptoflavins with iodoacetamide, N-ethylmaleimide, methyl methanethiolsulfonate, H202, and mchloroperbenzoate, in both the free and protein-bound state. The results confirm the conclusions drawn from the studies with 6-SCN-flavins described above and from 6-N3-flavins [Massey, V., Ghisla, S., &amp; Yagi, K. (1986) Biochemistry (preceding paper in this issue)]. The spectral properties of the protein-bound 6-mercaptoflavin vary widely among the five proteins studied and show stabilization of the neutral flavin with flavodoxin and riboflavin-binding protein and of the anionic species by Old Yellow Enzyme, lactate oxidase, and D-amino acid oxidase. In the case of the latter two enzymes, the stabilization appears to be due to interaction of the negatively charged flavin with a positively charged protein residue located near the flavin pyrimidine ring. This positively charged residue appears to be responsible also for the strong stabilization of the two-electron oxidation state of the mercaptoflavin as the 6-S-oxide. With the other flavoproteins studied this oxidation level is stabilized as the 6-sulfenic acid or 6-sulfenate.</dcterms:abstract>
    <dcterms:bibliographicCitation>First publ. in: Biochemistry 25 (1986), 24, pp. 8103-8112</dcterms:bibliographicCitation>
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    <dc:contributor>Massey, Vincent</dc:contributor>
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    <dc:contributor>Yagi, Kunio</dc:contributor>
    <dc:creator>Yagi, Kunio</dc:creator>
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    <dcterms:title>6-Thiocyanatoflavins and 6-mercaptoflavins as active-site probes of flavoproteins</dcterms:title>
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