T. Macneil et al., DELETION ANALYSIS OF THE AVERMECTIN BIOSYNTHETIC GENES OF STREPTOMYCES-AVERMITILIS BY GENE-CLUSTER DISPLACEMENT, Journal of bacteriology, 175(9), 1993, pp. 2552-2563
Streptomyces avermitilis produces a group of glycosylated, methylated
macrocyclic lactones, the avermectins, which have potent anthelmintic
activity. A homologous recombination,strategy termed gene cluster disp
lacement was used to construct Neo(r) deletion strains with defined en
dpoints and to clone the corresponding complementary DNA encoding func
tions for avermectin biosynthesis (avr). Thirty-five unique deletions
of 0.5 to >100 kb over a continuous 150-kb region were introduced into
S. avermitilis. Analysis of the avermectin phenotypes of tbe deletion
-containing strains defined the extent and ends of the 95-kb avr gene
cluster, identified a regulatory region, and mapped several avr functi
ons. A 60-kb region in the central portion determines the synthesis of
the macrolide ring. A 13-kb region at one end of the cluster is respo
nsible for synthesis and attachment of oleandrose disaccharide. A 10-k
b region at the other end has functions for positive regulation and C-
5 O methylation. Physical analysis of the deletions and of in vivo-clo
ned fragments refined a 130-kb physical map of the avr gene cluster re
gion.