Tubulin binding of two 1-deaza-7,8-dihydropteridines with different biological properties: Enantiomers NSC 613862 (S)-(-) and NSC 613863 (R)-(+)

• Published in: Biochemistry (Easton) Vol. 32; no. 40; pp. 10675 - 10682
• Main Authors: Leynadier, Daniel, Peyrot, Vincent, Sarrazin, Marcel, Briand, Claudette, Andreu, Jose Manuel, Rener, Gregory A, Temple, Caroll
• Format: Journal Article
• Language: English
• Published: Washington, DC American Chemical Society 12.10.1993
• Subjects: Animals; Antineoplastic agents; Antineoplastic Agents - chemistry; Antineoplastic Agents - metabolism; Biological and medical sciences; Brain; Cattle; General aspects; Kinetics; Medical sciences; Molecular Structure; Pharmacology. Drug treatments; Protein Binding; Pyrazines - chemistry; Pyrazines - metabolism; Spectrometry, Fluorescence; Spectrophotometry; Stereoisomerism; Structure-Activity Relationship; Tubulin - chemistry; Tubulin - isolation & purification; Tubulin - metabolism; Antineoplastic agent; Biological fixation; Tubulin; High affinity site; Enantiomer; Contractile protein; Enantiospecificity; Mechanism of action; In vitro; Isomer
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi00091a018

Several fluorescence properties of two enantiomers, NSC 613862 (S)-(-) and NSC 613863 (R)-(+), have been compared. Even though the two isomers showed the same fluorescence behavior in solution in different solvents, drastic differences were observed after binding to purified calf brain tubulin. Binding measurements for the two compounds were performed both by fluorescence spectroscopy and by column gel permeation, a direct method of measurement. For both isomers, the binding was characterized by the presence of one high-affinity binding site with an apparent association constant of (3.2 +/- 0.5) x 10(6) M-1 and (4.1 +/- 0.9) x 10(6) M-1 for the R- and S-isomer, respectively, and by several low-affinity sites. Both isomers were also shown to induce GTPase activity in tubulin. The high-affinity binding site seems to be the same for the two isomers. Moreover, fluorescence competition experiments suggest at least a partial overlap of the colchicine and podophyllotoxin site. To explain the differences in fluorescence behavior after binding to tubulin, we hypothesize that the R-isomer is positioned differently in its binding locus as compared with the S-isomer.