O

SCoA

OH O

OH O

SNAC

SNAC

SNAC

SNAC

6-deoxy-erythronolide B

Fig. 35.6. Incorporation by feeding of alternative diketide precursors into the erythromycin pathway results in novel variants of erythromycin A.

6-deoxy-erythronolide B

Fig. 35.6. Incorporation by feeding of alternative diketide precursors into the erythromycin pathway results in novel variants of erythromycin A.

6-deoxyerythronylide B (see Fig. 35.2A). This aglycon unit is processed by downstream enzymes that oxidize the backbone and append deoxysugars to give eryth-romycin A.

To explore the potential of combinatorial biosynthesis with type I modular PKSs, the DEBS system has been manipulated in several ways to produce combinatorial biosynthetic libraries. At the entry of the pathway, new biosynthetic products can be made either by utilizing alternate diketides (Fig. 35.6), which has also been possible with type II PKSs [43], or by changing the loading module [44]. Propionyl CoA is the natural starter unit, but replacing the DEBS loading module with the avermectin PKS loading module, which demonstrates broader specificity, results in the production of new polyketides (Fig. 35.7) [44]. In the pathway, combinatorial libraries have been produced by (1) altering the extension of the chain by replacing the native acyl transferase units in modules with non-native acyl transferase domains [45], (2) altering the state of reduction of the ketone at each chain position through changes in the reduction domains in each module [46], (3) altering the stereochemistry at tertiary and quaternary carbons through alteration in the appropriate domains [47, 48], or (4) feeding substrates that can be utilized by downstream modules [49, 50]. Additionally, diverse products can be made by lengthening or shortening the total length of the chain through adding or removing modules [5153].

Most DEBS-derived libraries to date, while presenting exciting advances in combinatorial biosynthesis, have been small by combinatorial library standards [54], since genetic manipulation of the very large type I modular PKS systems can be cumbersome. A clever approach that should allow for easier manipulation and larger combinatorial libraries is to utilize multiple PKS gene-containing plasmids

,SCoA

Propionyl-CoA

Was this article helpful?

0 0

Post a comment