Some polyketide-derived bioactive compounds contain sugars attached to the aglycone core, and these sugars often enhance or impart specific biological activity to the molecule. Mycinamicin II, a 16-member macrolide antibiotic produced by Micromonospora griseorubida A11725, contains a branched lactone and two different deoxyhexose sugars, D-desosamine and D-mycinose, at the C-5 and C-21 positions, respectively. We previously engineered an expression plasmid pSETmycinose containing the D-mycinose biosynthesis genes from M. griseorubida A11725. This plasmid was introduced into Micromonospora sp. FERM BP-1076 cells, which produce the 16-membered macrolide antibiotic izenamicin. The resulting engineered strain TPMA0041 produced 23-O-mycinosyl-20-deoxy-izenamicin B(1) and 22-O-mycinosyl-izenamicin B(2). 23-O-mycinosyl-20-deoxy-izenamicin B(1) has been produced by the engineered strain M. rosaria TPMA0001 containing pSETmycinose as 23-O-mycinosyl-20-deoxo-20-dihydro-12,13-deepoxyrosamicin (=IZI) in our recent study, and 22-O-mycinosyl-izenamicin B(2) has previously been synthesized as a macrolide antibiotic TMC-016 with strong antibacterial activity. The production of 22-O-mycinosyl-izenamicin B(2) (=TMC-016) was increased when propionate, a precursor of methylmalonyl-CoA, was added to the culture broth.