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J Bacteriol. 1969 November; 100(2): 557-564
Copyright © 1969 American Society for Microbiology. All Rights Reserved.

Characterization of the Biotin Transport System in Saccharomyces cerevisiae¹

^a Department of Microbiology, University of Cincinnati College of Medicine, Cincinnati, Ohio 45219

ABSTRACT

The characteristics of the biotin transport mechanism of Saccharomyces cerevisiae were investigated in nonproliferating cells. Microbiological and radioisotope assays were employed to measure biotin uptake. The vitamin existed intracellularly in both free and bound forms. Free biotin was extracted by boiling water. Chromatography of the free extract showed it to consist entirely of d-biotin. Cellular bound biotin was released by treating cells with 6 N H₂SO₄. The rate of biotin uptake was linear with time for 10 min, reaching a maximum at about 20 min followed by a gradual loss of accumulated free vitamin from the cells. Biotin was not degraded or converted to vitamers during uptake. Transport was temperature- and pH-dependent, optimum conditions for uptake being 30 C and pH 4.0. Glucose markedly stimulated biotin transport. In its presence, large intracellular free-biotin concentration gradients were established. Iodoacetate inhibited the glucose stimulation of biotin uptake. The rate of vitamin transport increased in a linear fashion with increasing cell mass. The transport system was saturated with increasing concentrations of the vitamin. The apparent K_m for uptake was 3.23 x 10^–7M. Uptake of radioactive biotin was inhibited by unlabeled biotin and a number of analogues including homobiotin, desthiobiotin, oxybiotin, norbiotin, and biotin sulfone. Proline, hydroxyproline, and 7,8-diaminopelargonic acid did not inhibit uptake. Unlabeled biotin and desthiobiotin exchanged with accumulated intracellular ¹⁴C-biotin, whereas hydroxyproline did not.

¹ Presented in part at the 68th Annual Meeting of the American Society for Microbiology, Detroit, Mich., 5-10 May 1968. Taken from a dissertation submitted by the senior author in partial fulfillment of the requirements for the Ph.D. degree in Microbiology.

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