Studies on the mode of antifungal action of pradimicin antibiotics. III. Spectrophotometric sequence analysis of the ternary complex formation of BMY-28864 with D-mannopyranoside and calcium.

Abstract

Sequence of reactions in the process of ternary complex formation of BMY-28864 with D-mannopyranosideand calcium was spectrophotometrically determined nnder more strict analytical conditions using metal-free preparations of sugars and the pradimicin derivative at a bandpass slit width of 1 nm. In the first phase of ternary complex formation, BMY-28864stereospecifically recognized and bound to D-mannopyranoside in the absence of calcium, which was revealed by a visible absorption maximumshift of ca. 8 nm. Subsequently, the BMY-28864-D-mannopyranoside conjugate reacted with calcium to yield the ternary complex, which was detected by an additional visible absorption maximumshift of ca. 8 nm. Whenthe three components were mixed at the same time, both phases simultaneously occurred to produce the ternary complex which was accompanied by a visible absorption maximumshift of 16 nm in total. Based on this two-phased reaction sequence, the mechanism of ternary complex formation of BMY-28864with D-mannopyranoside and calcium was reexamined in details. Terminal D-mannopyranoside was confirmed to be essential as BMY-28864-specific sugar receptor by in vitro analysis and animal cell experiments. While calcium, strontium and cadmium behaved similar in the in vitro ternary complex formation, the yeast and animal cell experiments showed that only calcium played a dual role as a base in the ternary complex formation and as an effector in physiological disturbances leading to cell death.In previous papers1~6), the antifungal activities of pradimicin derivatives were positively correlated with their lectin-mimic ternary complex formation by validating the essential roles of the C-18 carboxyl group and the C-5 thomosamine moiety of pradimicin derivatives. In consequence, characterization of the sugar-recognizing ability of pradimicin derivatives in the light of lectin biochemistry using highly-purified metal-free sugar and pradimicin preparations was considered to be informative and indispensable, as calcium and metal impurities in reagents probably interact or compete with the calcium componentof the ternary complex.The findings described in the preceding paper6) that BMS-184497 or BMY-28864methyl ester still retains as ability to showa visible absorption maximumshift of ca. 8nmin spite of the absence of the free C-18 carboxyl group; and that the breadth of the visible absorption peak shift depends on the status of the C-18 carboxyl group {ca. 8nm for the methyl ester and ca. 16nm for the free acid) allowed the hypothesis that the process of ternary complex formation might be divided into the sugar-recognization phase and the calcium salt formation phase. Therefore, the authors have attempted to elucidate the sequence of events in the visible absorption peak shift which seemed to be parallel with the process of ternary Correspondence should be addressed to Jun Okumura, Bristol-Myers

Pradimicin S, a Highly Soluble Nonpeptidic Small-Size Carbohydrate-Binding Antibiotic, Is an Anti-HIV Drug Lead for both Microbicidal and Systemic Use

“…Since no N-glycan deletions have ever been observed in gp41 of PRM-A-resistant (8) and PRM-Sresistant (this study) virus strains, this binding to gp41 may most likely not be directly related to the eventual antiviral activity of the pradimicins. The pradimicins have been suggested to be physiologically active as a (dimeric) complex with Ca 2ϩ (17,29,30). Experimental evidence has indeed been provided that in the presence of PRM-A, (␣-1,2)mannose oligomers, and Ca 2ϩ , a ternary complex is formed that consists of one Ca 2ϩ ion, two pradimicin-A molecules and four (␣-1,2)mannose oligomers (17,29,30).…”

“…The pradimicins have been suggested to be physiologically active as a (dimeric) complex with Ca 2ϩ (17,29,30). Experimental evidence has indeed been provided that in the presence of PRM-A, (␣-1,2)mannose oligomers, and Ca 2ϩ , a ternary complex is formed that consists of one Ca 2ϩ ion, two pradimicin-A molecules and four (␣-1,2)mannose oligomers (17,29,30). In the absence of Ca 2ϩ , PRM-A and PRM-S are unable to bind to HIV-1 gp120 in SPR experiments.…”

Pradimicin A, a d-mannose-binding antibiotic, binds pyranosides of l-fucose and l-galactose in a calcium-sensitive manner

“…10 These sugars have an arrangement similar to the 2-, 3-, and 4-hydroxyl groups of D-Man (Fig. 3), and are thus expected to bind to PRMs in the same manner as D-Man.…”

“…1), an artificial PRM derivative with improved water solubility, 8 revealed that PRMs and Ca 2+ ion form the binary [PRM/Ca 2+ ] complexes, which bind D-Man but not L-mannose, 2-, 3-, and 4-epimers of D-Man (D-glucose, D-altrose, and D-talose, respectively), D-mannosamine, and N-acetyl-D-mannosamine. 9,10 This sugar-binding profile led to the assumption that PRMs recognize the 2-, 3-, and 4-hydroxyl groups of D-Man.…”

Simian Immunodeficiency Virus Is Susceptible to Inhibition by Carbohydrate-Binding Agents in a Manner Similar to That of HIV: Implications for Further Preclinical Drug Development

“…It specifically binds to the terminal D-mannose pyranosides of ␣(1,2)-mannose oligomers. It requires the presence of Ca 2ϩ , which is thought to bind two PRM-A molecules through the free carboxylic acid group present on each molecule (Ueki et al, 1993).…”