Pradimicin L

The biosynthetic pathway of pradimicin S (PRM-S) was investigated by using sinefungin and bioconversion experiments with aglycones of pradimicin A (PRM-A) and Actinomadura spinosa AA0851, a PRM-S producer. Addition of sinefungin to the strain inhibited the formation of 11-O-demethyl-7-O-methylpradinone II (11dM-7M-PNII) as also determined to occur with its addition to the PRM-A producer. In feeding PRM-A aglycone and its analogs to the strain early in PRM-S biosynthesis, good identifications of bioconverted products were obtained by frit-FAB LC/MS as follows: 11-O-demethylpradinone II (11dM-PNII), 11dM-7M-PNII, 11-O-demethylpradinone I (11dM-PNI), 11-O-demethylpradimicinone I (11dM-PMNI) and pradimicinone I (PMNI) were converted to PRM-S. Pradimicin B (PRM-B) and pradimicin L (PRM-L) were converted to PRMs-L and -S and PRM-S, respectively. A biosynthetic pathway for PRM-S is proposed.

Pradimicin is an antifungal antibiotic which induces apoptosis like cell death in the yeast Saccharomyces cerevisiae. Pradimicin-resistant mutants were isolated from the S. cerevisiae and the mutation points were analyzed. A point mutation of YPD1 that led to a substitution of the 74th glycine (Gly74) to cysteine (Cys) was identified in a mutant strain NH1. In S. cerevisiae, Ypd1 transfers a phosphoryl group from the sensor kinase Slnl to the response regulator Sskl which regulates a downstream MAP kinase in response to hyperosmotic stress. Gly74 is located in a three-residue reverse turn domain that connects two alpha-helices, one of which contains a histidine residue which is phosphorylated. In the reverse turn, glycine (relative position +10 to the active-site histidine) is highly conserved in Ypd1 and other histidine-containing phosphotransfer proteins. It was therefore suggested that the substitution of Gly74 to Cys altered the Ypd1 structure, which resulted in the resistance to pradimicin.

Pradimicin S was isolated from the culture filtrate of Actinomadura spinosa AA0851. NMR and MS analyses proved that pradimicin S is the 3'-O-(3''-O-sulfo-beta-D-glucopyranosyl) analog of pradimicin A, a new member of the pradimicin family of antibiotics. Stereochemical assignment was made by correlating pradimicin S with pradimicin L.